P S Kewate et al.

/ Journal of Pharmacy Research 2015,9(3),182-189

Research Article
ISSN: 0974-6943

Available online through

www.jpronline.info

In silico enzyme function prediction in hypothetical proteins of Mycobacterium bovis AF2122/97

P S Kewate1, R C Urade1, D G Gore2*, M A Soni1 and A P Kopulwar1
Priyadarshini Institute of Engineering and Technology (Department of Biotechnology),
Mouza Shivangaon, Behind CRPF Campus, Hingna Road, Nagpur 440019.
2
Sai Bioinfosys Institute of Bioinformatics Research, Plot No. 271 Raghuji Nagar, Nagpur (Maharashtra), India.440009
1

Received on:28-01-2015; Revised on: 19-02-2015; Accepted on:22-03-2015

ABSTRACT
Background: Mycobacterium bovis linked to cause bovine tuberculosis has been investigated and functionally annotated for several
hypothetical proteins ability of enzyme activity as per the conserved domain present. Methods: As M. bovis genome codes in total 25%
hypothetical proteins, we used CDD BLAST, INTERPRSCAN, PFAM and CATH domain search programs to find the enzyme functionality in
these proteins. Results and discussion: Programs reported 347 hypothetical proteins posses enzymatic activity as per the enzyme domain
conserved in the sequences at variable confidence levels. Conclusion: These proteins could be the markers, to investigate about their fate
in the metabolism of M. bovis.
KEYWORDS: Bioinformatics, Hypothetical proteins, Enzyme conserved domain.
INTRODUCTION
Mycobacterium bovis (M. bovis) is the etiological agent of bovine
tuberculosis and a close relative of Mycobacterium tuberculosis (M.
tuberculosis) in the family of M. tuberculosis complex (MTBC), which
consists of M. tuberculosis, M. bovis, M. bovis BCG, M. africanum,
M. canettii, M. microti, M. caprae, and M. pinnipedii. The M. bovis
mostly infects cattle (Bos taurus), but it can also infect other mammalians including humans.1, 2 BCG, an attenuated strain of M. bovis, has
been used in many countries or territories as a prophylactic vaccine
for disease tuberculosis (TB), for more than 90 years, although a
considerable controversy related to its efficacy. The true efficiency
of BCG has not so easy to understand due to many experimental
variables.3 The BCG vaccine undoubtedly provides protection against
childhood disseminated form of TB including TB meningitis. However its efficacy against pulmonary TB in adults has been reported to
give variable results.4An estimated 8.6 million new cases and 1.3 million deaths due to tuberculosis occur every year.5 Almost all cases of
tuberculosis are caused by M. tuberculosis, and share of M. bovis is
less than 1.4 percent of all pulmonary tuberculosis cases outside of
Africa. Though, in Africa, M. bovis accounts for approximately 2.8
percent of cases of pulmonary tuberculosis, for a crude incidence of
7 cases per 100,000 populations.6 In view, a study has been conducted to computationally assign the enzyme function on the hypo-

thetical protein islands, which could be useful to elucidate further

about hidden potential of M. bovis.
MATERIALS AND METHODS
Data Collection
The hypothetical proteins of Mycobacterium bovis AF2122/97 (mbo)
7
have been gathered from the website www.genome.jp/kegg/ available in the FASTA format.
Search for Conserved Sequences
The 468 hypothetical proteins of M. bovis were screened for the presence of enzyme conserved domain in the query sequences by giving
them as input to the four conserved domain searching programs.
These programs remain linked with the protein databases and assist
in annotating the functional homology of the query sequence with
the best scored protein family. The programs used as:A) CDD-BLAST (http://www.ncbi.nlm.nih.gov/BLAST/) 8, 9, 10
B) INTERPROSCAN (http://www.abi.ac.uk/interpro) 11
C) PFAM (http://www.pfam.sanger.ac.uk/)12
D) CATH (http://www.cathdb.info/)13

Functional Categorization
The hypothetical proteins possessing enzyme domains as per domain searching programs were categorized in percent confidence levels using following parameters:1. Four tools indicate the same enzyme functions then confidence
level set as 100 percent.
Journal of Pharmacy Research Vol.9 Issue 3.March 2015
182-189

*Corresponding author.
D G Gore
Sai Bioinfosys Institute of Bioinformatics Research,
Plot No. 271 Raghuji Nagar,
Nagpur (Maharashtra), India.440009

P S Kewate et al. / Journal of Pharmacy Research 2015,9(3),182-189

2. Three tools indicate the same enzyme functions and one showing
different function then the confidence level set as 75 percent.
3. Two tools indicate the same enzyme functions and two showing
different functions then the confidence level set as 50 percent.
4. One tool indicates the enzyme function and three tools showing
different functions then the confidence level set as 25 percent.
OBSERVATIONS AND RESULTS
Functional Annotation to the Hypothetical Proteins
The search for domains in the hypothetical protein have been done
with CDD-BLAST, CATH, INTERPROSCAN and PFAM, all these

web tools has accepted the FASTA format sequence of hypothetical

protein and analyzed the hidden enzymatic conserved domains in all
347 hypothetical proteins and classified them into enzyme family with
given confidence level as follows 100% for 135 proteins, 75% for 119,
50% for 52, 25% for 45 as represented in Table 01. The four programs
result suggested that even though, the genome posses 25 % genes
for the hypothetical proteins those contain the conserved domain as
347 hypothetical proteins showed the defined presence of enzyme
function. Based on the % confidence it has been noteworthy that
proteins with 100% confidence could be first choice for In Vivo analysis which will assist in determining the fate of these well annotated
hypothetical proteins in the metabolism of M. bovis.

Table 1: Enzyme funtion predition in the hypothetial proteins of M. bovis

KEGG NO Interproscan5

PFAM

CDDBLAST

Mb0021
Mb0062
Mb0065
Mb0076
Mb0083
Mb0101
Mb0103
Mb0105
Mb0107
Mb0114
Mb0120
Mb0126
Mb0132
Mb0143
Mb0146
Mb0147
Mb0150
Mb0151
Mb0168

Aldolase-type TIM barrel

esterase
PIN
Amidohydrolase
FMN binding
Long-chain fatty acyl-CoA thioesterase,
Acyl carrier protein
Cytochrome c oxidase
NAD binding domain
Peptidase
L,D-transpeptidase
FMN-binding
Protein Kinase
Polyketide cyclase
Polyketide cyclase SnoaL
DNA glycosylase
Leucine carboxyl methyltransferase
Leucine carboxyl methyltransferase
Thioesterase superfamily

Nitronate monooxygenase
2-Nitropropane dioxygenase
glycohydro6
glycohydro6
PIN
PIN
amidohydro1
met dependent hydro A
pyridox ox2
Pyridox ox2
FcoT
FcoT-like thioesterase domain
Phosphopantetheine
PP-binding
Cytochrome c oxidase
Cytochrome c oxidase
Cyclic nucleotide binding
glycerate dehydrogenases
Rhomboid
Rhomboid
L,D-transpeptidase
L,D-transpeptidase
Pyridox_oxidase
PPOX
Phosphotransferasr
Ble
Snoal like
SnoaL
Snoal like
Ketosteroid isomerase
NO
3-methyladenine DNA glycosylase
Leucine carboxyl methyltransferase O-Methyltransferase
Leucine carboxyl methyltransferase methyltransferase
Thioesterase
Thioesterase

Mb0169
Mb0173
Mb0174
Mb0187
Mb0191
Mb0196
Mb0197
Mb0222
Mb0227
Mb0230
Mb0234
Mb0247
Mb0269
Mb0270

Streptomyces cyclase
Permease
Permease
Pirin
NO
Major facilitator
L,D-transpeptidas
Thioesterase superfamily
O-acyltransferase,
Glyco tran
PIN
Maoc
Allophanate hydrolase
Allophanate hydrolase
DNA primase
Anhydrolase
lyoxalase
Leucine carboxyl methyltransferase
nucleoside triphosphate hydrolase
nucleoside triphosphate hydrolase
Ribbon helix protein
PIN
Phosphatidic acid phosphatase
LD transpeptidase catalytic
NO
no
Zinc permease
putative decaetylase LMBE
methyltransferase
methyltransferase
HTH Type,Tetr-N

Polyketide_cyc
Srpbcc domain
Permease
Ttg
Permease
Ttg2B
Pirin
Pirin
NO
putative metallohydrolase
Major Facilitator
Arabinose efflux permease
LD transpeptidase
L,D-transpeptidase
NO
FkbR
WES_acyltransf
acyltransf
Glyco_trans_
Glycosyltransferas
PIN
PIN Mrv
MaoC_dehydratas
Maoc dehydrates
Allophanate hydrolase
Allophanate hydrolase
Allophanate hydrolase
Allophanate hydrolase
DNA_primase
Ligd
Anhydrolas
Anhydrolase
Glyoxalase
Sphingobium chlorophenolicum
PE
PE
Leucine carboxyl methyltransferase o methyltransferase
FtsK
FtsK
Abhydrolase
Abbhydrolase6
RHH1
PHA
PIN
PIN
PAP2
PAP2
LD transpeptidase catalytic
NO
Snoal like
snoal like
NO
NO
NO
NO
PIGL-L
PIG
methyltransferase
AT4G
methyltransferase
Adomet-Mtases

Mb0278
Mb0280
Mb0285
Mb0289
Mb0292
Mb0301
Mb0308
Mb0309
Mb0315
Mb0316
Mb0318
Mb0322
Mb0328
Mb0331
Mb0333
Mb0336

CATH

NO
75
NO
75
smg6
100
no
75
NO
75
NO
75
NO
75
NO
75
Adenosylhomocysteinase
100
Rhomboid
100
Erfk Family like
75
FMN binding protein
100
NO
50
Steroid Delta-isomerase
100
NO
75
N-glycosylase/DNA lyase
75
Putative S-adenosyl-L-methionine100
Putative S-adenosyl-L-methionine
100
1,4-dihydroxy-2-naphthoyl100
CoA hydrolase
NO
75
NO
50
NO
50
Quercetin 2,3-dioxygenase
100
NO
25
Sugar efflux transporter B
25
ErfK
100
ribosomal
75
NO
75
NO
75
NO
75
Nodulation like
100
NO
75
NO
75
NO
50
Anhydrolase
100
4 hydroxyphenylpyruvate dioxygenase 100
NO
25
Putative S-adenosyl-L-methionine
100
FtsK
25
NO
50
NO
25
SMG6 PROTEIN
75
Lipid phosphatidic phosphatase
50
NO
25
Bile acid 7 alpha dehydratase
75
NO
25
NO
25
1D Myo-inositol
50
AT4G
50
NO
75

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P S Kewate et al. / Journal of Pharmacy Research 2015,9(3),182-189

KEGG NO Interproscan5
Mb0339
Mb0340
Mb0347
Mb0353
Mb0357
Mb0365
Mb0366
Mb0368
Mb0372
Mb0373
Mb0375
Mb0378
Mb0379
Mb0383
Mb0390
Mb0392
Mb0396
Mb0399
Mb0404
Mb0410
Mb0420
Mb0432
Mb0438
Mb0442
Mb0449
Mb0452
Mb1098
Mb1102
Mb1107
Mb1110
Mb1113
Mb1116A
Mb1127
Mb1134
Mb1135
Mb1139
Mb1144
Mb1145
Mb1156
Mb1159
Mb1172
Mb1179
Mb1185
Mb1187
Mb1188
Mb1189
Mb1216
Mb1223
Mb1224
Mb1226
Mb1236
Mb1237
Mb1241
Mb1248
Mb1254
Mb1259
Mb1263
Mb1278
mb1302
Mb1292
Mb1296
Mb1300
Mb1308
Mb1309
Mb1315
Mb1319

Polyketide cyclase snoal like

NO
MobA-like NTP transferase
NO
Thioesterase
Peptidase
NO
Six-hairpin glycosidase-like
zeta toxin
VWA
NTP transferase
Xdhc and coxl
Xdhc and coxl
Dodein
oxidoreductase
Rhodanese
HNH NUCLEASE
NO
Antinomycetalese membrane
NO
NO
aceyl coA
Lon protease
NO
Putative metal dependent hydrolase
Amino oxidase
Rhodanese Homolog
Abhydrolase_
SGNH_hydrolase
RDD
NO
thioredoxin
Decaprenyl diphosphate synthase
PemK
Carboxylesterase
Carboxylesterase
NO
PIN
Adenylate &Guanylate
cyclase catalyti
NO
Isoprenylcysteine
carboxyl methyltransfe
S-adenosylmethionine
-dependent methyltransferase
HNH nucleases;
Pyridoxamine 5'-phosphate oxidase
HhH-GPD
Pistil-specific extensin-like protein
Helicase superfamily
c-terminal domain
Alpha/beta hydrolase
Alpha/Beta hydrolase
Abhydrolase
Tetratricopeptide
Cytokinin riboside
5'-monophosphate
phosphoribohydrolase LOG
Cytokinin riboside
5'-monophosphate
phosphoribohydrolase LOG
Xaa-Pro dipeptidyl-peptidas
Phospholipid methyltransferase
NO
Transglycosylase
PIN
Uracil-DNA glycosylase
FAD binding domain
Deazaflavin-dependent nitroreductas
Peptidase_
phosphoesterase domain
AAA+ATPase domain
Carbonic anhydrase (IPR001765)
Bacteriophage P2,
GpX (IPR008861)

PFAM

CDDBLAST

MDMPIC
DinB2
Snoal like
snoal like
NO
NO
MobA-like NTP transferase domain Glycosyltransferase family
NO
NO
NO
NO
Peptidase family
Peptidase
NO
Glyco_tranf_GTA_
Glycosyl hydrolase
Predicted glycosyl hydrolase
Zeta toxin
AAA domain
VWA
vWFA
NTP transferase
glycosyltransferase
XdhC and CoxI
XdhC Rossmann
XdhC and CoxI
XdhC and CoxI
NO
Proline-rich
oxidoreductase
reductase
Rhodanese
Rhodanese Homolog
NO
HNH endonuclease
NO
NO
Mycobacterium membrane
Mycobacterium membrane protein
Alpha/beta hydrolase
Alpha/beta hydrolase
NO
NO
NO
NO
Lon protease
Lon protease
NO
NO
Rska
rskA
Rhodanese Homolog
Rhodanese Homolog
Abhydrolase_
Abhydrolase_
GDSL-like Lipase/
SGNH_hydrolase
RDD
RDD
NO
SIS
thioredoxin
thioredoxin
diphosphate synthase
undecaprenyl diphosphate synthase
PemK
MazF
Carboxylesterase
Carboxylesterase
Carboxylesterase
Esterases and lipases
NO
YbjN
PIN
PIN
Adenylate& Guanylate
Adenylate & Guanylate
cyclase catalyti
cyclase
NO
acyltransferase
Isoprenylcysteine
methyltransferase
carboxyl methyltransfe
S-adenosylmethionine
SAM methyltransferase
-dependent methyltransferase
HNH nucleases;
HNH nucleases;
Pyridoxamine 5'-phosphate oxidase PPOX
HhH-GPD
HhH-GPD
NO
alpha/beta subunit
Helicase superfamily
Helicase
c-terminal domain
Alpha/beta hydrolase
Alpha/beta hydrolase
hydrolase
Putative lysophospholipase;
Abhydrolase
Abhydrolase
AAA16
AAA16
Lysine_decarbox
TIGR

CATH

NO
50
NO
50
NO
25
NO
50
NO
25
NO
25
NO
50
NO
50
NO
75
putativ
50
collagen aipha3
25
NO
75
NO
75
NO
50
NO
25
reductase
100
Calcium sensing receptor,
75
NO
25
NO
25
NO
50
embryogenic ass.
75
NO
25
NO
25
NO
50
NO
25
NO
25
tRNA sulfurtransferase
100
NO
75
Conserved exported
75
RDD
100
NO
25
Spermatogenesis
75
Undecaprenyl pyrophosphate synthase 100
25
NO
75
NO
75
NO
25
PIN
100
Adenylate cyclas
100
NO
NO
MPBQ/MSBQ methyltransferas

100

NO
FMN-binding protein
N-glycosylase/DNA lyase
NO
ATP-dependent DNA
helicase uvsW
Haloalkane dehalogenase
NO
Put ative uncharacterized protein
ATP-dependent protease
Cytokinin riboside
5'-monophosphate

75
100
100
25
100

Lysine_decarbox

TIGR

Cytokinin riboside
5'-monophosphate

Xaa-Pro dipeptidyl-peptidas
Phospholipid methyltransferase
NO
Transglycosylase
PIN
Uracil-DNA glycosylase
FAD binding domain
NO
Peptidase_
Calcineurin-like phosphoesterase
AAA domain
Carbonic anhydrase
LysM domain

Xaa-Pro dipeptidyl-peptidas
methyltransferase
PRK
Transglycosylase
PIN
Uracil-DNA glycosylase
hydroxylase
NO
Peptidase_
DNA repair exonuclease
AAA domain
Carbonic anhydrases (CA
hydrolase

Xaa-Pro dipeptidyl-peptidas
NO
NO
Transglycosylase
PIN
Uracil-DNA glycosylase
NO
NO
NO
NO
NO
NO
NO

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75

100
75
75
50
75
75
100
75
25
100
100
100
75
25
75
75
75
75
50

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P S Kewate et al. / Journal of Pharmacy Research 2015,9(3),182-189

KEGG NO Interproscan5
Mb1332
Mb1333
Mb1335
Mb1345
Mb1346
Mb1347

PFAM

CDDBLAST

Modification methylase
Methyltransferase domain
SAM methyltransferases
HemK (IPR004556)
NO
Telomere recombination
tRNA threonylcarbamoyl adenosine
ATP synthase protein I (IPR005598)
ATP synthase I chain
NO
Transposase
Transposase
Transposase
Transposase
Helix-turn-helix domain of
transposase
transposase family ISL3
ATP:cob(I)alamin adenosyltransferase
Cobalamin adenosyltransferase
cob(I)alamin adenosyltransferase

Mb1355
Mb1357

Endonuclease NucS (IPR002793)

MethylmalonylCoA epimerase

NO
Glyoxalase/
Bleomycin resistance protein

Mb1369

NO

Prokaryotic homologs
of the JAB domain

isopeptidase activity

Mb1372

Peptidase S54, rhomboid domain

Rhomboid family

Rhomboid family

Mb1374

Beta-lactamase-like

Mb1382

Acyl-CoA N-acyltransferase

Mb1392

Diguanylate phosphodiesterase

EAL domain

EAL domain

Mb1399

Protein phosphatase
2C (PP2C)-like domain

PAS fold

Sulphate Transporter
and Anti-Sigma factor

Mb1401

RelA/SpoT

Region found in
RelA / SpoT proteins

NT_Rel-Spo_like[cd05399],
Nucleotidyltransferase (NT)
domain of RelA- and SpoT-like pp
Gpp synthetases and hydrolases

Mb1402

Beta-lactamase/transpeptidase-like

Beta-lactamase

Beta-lactamase

Mb1405
Mb1406

Fatty acid desaturase, type 1

Polyketide synthase, type III

Fatty acid desaturase

Chalcone and stilbene synthases

Membrane-FADSChalcone and stilbene synthases

Mb1413
Mb1432

HNH nucleases
VapC family

NO
PIN domain

HNH nucleases
PIN domain of ribonucleases

Mb1433

Ribbon-helix-helix

Mb1448

Alanine racemase,
N-terminal domain

Mb1449
Mb1456

D-serine dehydratase-like domain

ATPase, P-loop-containing

Mb1457

NO
MethylmalonylCoA epimerase

Beta-lactamase superfamily
Metal-dependent
domain
hydrolase
Acetyltransferase (GNAT) domain Acetyltransferase (GNAT) domain

Ribbon-helix-helix
protein, copG family
Alanine racemase,
N-terminal domain

Ribbon-helix
-helix protein
Type III Pyridoxal 5-phosphate
(PLP)-Dependent Enzymes,
Low Specificity D-Threonine
Aldolase-like
Putative serine dehydratase domain Predicted amino acid aldolase or racemase
P-loop ATPase protein family
P-loop ATPase protein family

CATH

NO

75

NO
NO
NO
NO

50
50
75
75

cob(I)alamin
adenosyltransferase
NO
4-hydroxyphenylpyruvate
dioxygenase -like
domain 1/2
COP9 signalosome
complex subunit 5
-like domain
Rhomboid protein 2
-like domain
Hydroxyacylglutathione
hydrolases-like domain
Mycothiol
acetyltransferase -like
domain 1/2/3
Signaling protein
ykoW -like domain
Phosphoserine
phosphatase rsbP
-like domain
GTP pyrophosphokinase
-like domain

100

Glutaminase
-like domain
NO
Chalcone synthase
-like domain 1
NO
SMG6 protein
-like domain
Nickel-responsive
regulator -like domain
Alanine racemase
-like domain

100

Uncharacterised protein
family UPF0052

proteins similar to
B.subtilis YvcK

WhiA N-terminal LAGLID

WhiA LAGLIDADG-like domain

Mb1460

Acyltransferase, WS/DGAT/MGAT

Mb1463

Phospholipid/glycerol acyltransferase

Wax ester synthase-like

Acyl-CoA acyltra
ADG-like domain
Acyltransferase

Mb1468

L,D-transpeptidase
catalytic domain

L,D-transpeptidase
catalytic domain

Mb1474

Polyketide cyclase SnoaL-like domain

SnoaL-like domain

Mb1478

Polyketide cyclase/dehydrase

Polyketide cyclase /
dehydrase and lipid

Mb1517

Von Willebrand factor

type A (vWA) domain

Von Willebrand factor

type A (vWA) domain

Wax ester synthase-like

NO
Acyl-CoA acyltransferase
domain
Lysophospholipid Acyltransferases (LPLATs)
NO
of Glycerophospholipid Biosynthesis
IgD-like repeat domain
ErfK/YbiS/YcfS/YnhG
of mycobacterial L,Dfamily protein -like
transpeptidases
domain
SnoaL-like domain
Putative uncharacterized
protein -like domain
START/RHO_alpha_C/PITP/
NO
Bet_v1/CoxG/CalC (SRPBCC)
ligand-binding domain superfamily
Von Willebrand factor
Collagen alphatype A (vWA) domain
3(VI) chain -like domain

Mb1523

NfeD-like, C-terminal domain

NO

Journal of Pharmacy Research Vol.9 Issue 3.March 2015

75
100
100
100
25
100
100

75
100
50
50
25
100

NO
UPF0042 nucleotidebinding protein CMS1991
-like domain
2-phospho-L
-lactate transferase
-like domain
NO

LPPG:FO 2-phospho
-L-lactate transferase
CofD/UPF0052
Mb1458 Sporulation regulator WhiA

NfeD-like C-terminal,
partner-binding

25
100

Nodulation efficiency
protein D family protein
-like domain

75
75
50
75
75
75
100
75
75
100
75

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KEGG NO Interproscan5
Mb1526
Mb1532
Mb1539
Mb1541
Mb1543
NeuD
Mb1545
Mb1546

Mb1558
Mb1560
Mb1567
Mb1573

Mb1584
Mb1587
Mb1592
Mb1598
Mb1613
Mb1618
Mb1619
Mb1623
Mb1624
Mb1645
Mb1658
Mb1663
Mb1666
Mb1674
Mb1700
Mb1703
Mb1707
Mb1719
Mb1723
Mb1726
Mb1728
Mb1744
Mb1745
Mb1746
Mb1749

PFAM

CDDBLAST

CATH

Methylmalonyl-CoA mutase, Methylmalonyl-CoA mutase

Coenzyme B12-dependentNO
alpha/beta chain, catalytic
methylmalonyl coenzyme A
mRNA interferase
PemK-like protein
Growth inhibitor
mRNA interferase
PemK-like protein
MazF -like domain
Phytanoyl-CoA dioxygenase Phytanoyl-CoA dioxygenase (PhyH) Protein involved in biosynthesis
Phytanoyl-CoA hydroxylase -like domain 1/2
of mitomycin antibiotics/polyketide
fumonisin
Glycosyl hydrolase, fiveNO
Glycosyl hydrolase families
Alpha-L-arabinofuranosidase axhA -like domain
bladed beta-propellor domain
Sialic acid O-acyltransferase, Bacterial transferase hexapeptide
Putative Acyltransferase (AT), Left- UDP-3-O-acylglucosamine
handed parallel beta-Helix
N-acyltransferase -like domain
(LbH) domain
Nucleotide-diphospho
Glycosyl transferase family 2
Glycosyl transferase family 2
Polypeptide N-acetylgalactosaminyltransferase
sugar transferases
1 -like domain
DegT/DnrJ/EryC1/StrS
DegT/DnrJ/EryC1/StrS
Aspartate aminotransferase
NO
aminotransferase
aminotransferase
(AAT) superfamily
(fold type I) of pyridoxal
phosphate (PLP)
AhpD-like
Carboxymuconolactone
Carboxymuconolactone
NO
decarboxylase famil
decarboxylase family
Aldolase-type TIM barrel
Nitronate monooxygenase
Nitronate monooxygenase
NO
Pseudouridine synthase,
S4 domain
Pseudouridine synthase,
NO
RluC/RluD
RsuA/RluD family
Polyketide cyclase/dehydrase Polyketide cyclase
START/RHO_alpha_C/
Putative uncharacterized protein -like domain
/ dehydrase and lipid
PITP/Bet_v1/CoxG/CalC
(SRPBCC) ligand-binding
domain superfamily
Deazaflavin-dependent
NO
NO
NO
nitroreductase
VapC family
PIN domain
PIN domain
SMG6 protein -like domain
Acyltransferase 3
Acyltransferase family
Predicted acyltransferases
NO
[Lipid metabolism]
RNA ligase/cyclic
2'-5' RNA ligase superfamily
2'-5' RNA ligase superfamily
NO
nucleotide phosphodiesterase
HNH nucleases
NO
HNH nucleases
NO
Lipase, secreted
Secretory lipase
Secretory lipase
NO
NUDIX domain
NUDIX domain
ADP-ribose pyrophosphatase
NO
S-adenosyl-L-methionine Methyltransferase domain
S-adenosylmethionine-dependent
NO
-dependent
methyltransferases
methyltransferase-like
Deazaflavin-dependent
NO
deazaflavin-dependent oxidoreductase, NO
nitroreductase
nitroreductase family
Lysylphosphatidylglycerol NO
NO
NO
synthetase
NO
NO
Predicted RNA-binding protein,
RNA-binding protein AU-1 -like domain
Beta-lactamase-like
Metallo-beta-lactamase superfamily
Zn-dependent hydrolases, including Hydroxyacylglutathione hydrolase -like domain
associated with RNAses E/G family
glyoxylases
Putative esterase
Putative esterase
Predicted hydrolase of the
Pimelyl-[acyl-carrier protein]
alpha/beta superfamily
methyl ester -like domain
Adenylyl cyclase classAdenylate and Guanylate
cyclase homology domain
Adenylate cyclase -like domain
3/4/guanylyl cyclase
cyclase catalyti
Transglutaminase-like
Transglutaminase-like superfamily
Transglutaminase-like superfamily Putative uncharacterized protein -like domain
Thioredoxin-like fold
AhpC/TSA family
Peroxiredoxin (PRX)-like 2 family Nucleoredoxin -like domain 1/2
NO
ABC transporter, phosphonate,
ABC transporter, phosphonate,
NO
periplasmi
periplasmic substrate-binding
protein
methionyl-tRNA synthetase
NO
Thiamine pyrophosphokinase Thiamine pyrophosphokinase
Thiamine pyrophosphokinase
Thiamine pyrophosphokinase
C terminal
C terminal
C terminal
-like domain
NUDIX domain
NUDIX domain
ADP-ribose pyrophosphatase
RNA pyrophosphohydrolase -like domain
(ADPRase) catalyzes the
hydrolysis of ADP-ribose
HNH nucleases
NO
HNH nucleases
NO
Putative cyclase
Putative cyclase
Kynurenine formamidase
Kynurenine formamidase -like domain
[Amino acid transport
and metabolism]
RmlC-like cupin domain
Cupin domain
Cupin domain
Sucrose-binding protein -like domain 1/2
3-keto-5-aminohexanoate
NO
NO
NO
cleavage enzyme
VapC family
PIN domain
PIN domain of the
SMG6 protein -like domain
Pyrobaculum aerophilum Pae0151
and Pae2754 proteins and homologs

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100
100
75
100
100
75

75
75
75
75

25
100
75
50
50
75
75
75
50
25
50
100
100
100
75
100
50
50
100
100
50
100
100
25
100

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P S Kewate et al. / Journal of Pharmacy Research 2015,9(3),182-189

KEGG NO Interproscan5

PFAM

Mb1754

HxlR-like helixturn-helix

HxlR-like helix-turn-helix

Mb1756

Conserved hypothetical
protein CHP03083,
actinobacterial-type
Leucine carboxyl
methyltransferase

Mycothiol maleylpyruvate
isomerase N-ter

Mb1758

Leucine carboxyl methyltransferase

Mb1761

Thioredoxin-like fold

Mb1763
Mb1770
Mb1780
Mb1781

Chloramphenicol acetyltransferase
-like domain
PIN domain-like
Aromatic-ring hydroxylase-like
NO

Mb1785
Mb1791
Mb1794

Glycosyl transferases group 1

Glycosyl transferases group 1
Acyltransferase, WS/DGAT/MGAT acyltransferase
HNH endonuclease
HNH endonuclease

Mb1796

AhpD-like

Mb1798

PLP-binding barrel

Mb1799

Peptidase family M28

Peptidase family M28

Mb1812
Mb1822
Mb1823

Type VII secretion system EccCa

EspG family
YukD-like

Mb1826

CbxX/CfqX

FtsK/SpoIIIE family
EspG family
WXG100 protein secretion
system (Wss), p
ATPase

Mb1838
Mb1845

PPE family
Trypsin-like cysteine/
serine peptidase domain
Succinate dehydrogenase/fumarate
reductase flavoprotein,
catalytic domain
Thiamine pyrophosphate enzyme,
N-terminal TPP-binding domain

Mb1848
Mb1851

AhpC/TSA family
2-oxoacid dehydrogenases
acyltransferase
NO
FAD binding domain
NO

Carboxymuconolactone
decarboxylase
Alanine racemase, N-terminal
domain

CDDBLAST

O-Methyltransferase involved in
polyketide biosynthesis [Secondary
metabolites biosynthesis, ...
Peroxiredoxin (PRX)-like 1 family

NO

25

Putative S-adenosyl-L-methionine
-dependent -like domain

100

Glutathione peroxidase
-like domain
Dihydrolipoyllysine-residue
succinyltransferase -like domain
NO
NO
Inositol 2-dehydrogenase -like domain

2-oxoacid dehydrogenases
acyltransferase (catalytic domain)
PIN (PilT N terminus) domain
NAD(P)-binding Rossmann-like domain
Predicted dehydrogenases and related
proteins [General function prediction only]
Glycosyltransferase
NO
acyltransferase domain
NO
HNH endonuclease
NO
gamma-carboxymuconolactone
decarboxylase
Type III Pyridoxal 5-phosphate
(PLP)-Dependent Enzymes Similar
to D-Serine Dehydratase and
Zinc peptidases M18, M20,
M28, and M42
FtsK/SpoIIIE family
EspG family
type VII secretion integral membrane
protein EccD
The AAA+ (ATPases

100
100
50
75
50
75
75
75

NO

50

Alanine racemase -like domain

100

Cytosol aminopeptidase -like domain

100

FtsK -like domain

NO
NO

75
75
75

Tetratricopeptide repeat protein

30A -like domain 1/2/3
NO
Genome polyprotein -like domain 1/2

100

FAD binding domain

NAD(P)-binding Rossmann-like domain L-aspartate oxidase -like domain

100

Thiamine pyrophosphate (TPP) family,

1-deoxy-D-xylulose-5-phosphate
synthase -like domain 1/2/3
Bifunctional nuclease
X-Pro dipeptidyl-peptidase C-terminal
non-catalytic domain
Von Willebrand factor type A (vWA)
domain was originally found in
the blood coagulation ...
This cd contains two tandem repeats
of the cystathionine beta-synthase
(CBS pair) domains ...
CBS domain
Zn-dependent protease with
chaperone function
Thioesterase-like superfamily

1-deoxy-D-xylulose-5-phosphate
synthase -like domain 1/2/3

100

NO
Xaa-Pro dipeptidyl-peptidase
-like domain
Mediator of RNA polymerase II
transcription -like domain

75
100

KpsF protein -like domain 1/2

100

KpsF protein -like domain 1/2

Protease HtpX -like domain

100
100

1,4-dihydroxy-2-naphthoyl-CoA
hydrolase -like domain
NO
Putative uncharacterized
protein -like domain 2
NO
NO
NO
FMN-binding protein -like domain

100

Thiamine pyrophosphate
enzyme, N-termina

Mb1867

von Willebrand factor, type A

Mb1872

CBS domain

CBS domain

Mb1873
Mb1876

CBS domain
Peptidase family M48

CBS domain
Peptidase family M48

Mb1878

Thioesterase-like superfamily

Thioesterase-like superfamily

Mb1894
Mb1897

CAAX amino terminal protease

CoA-transferase family III

CAAX protease self-immunity

CoA-transferase family III

Bifunctional nuclease
X-Pro dipeptidyl-peptidase
(S15 family)
Bacterial extracellular
solute-binding p

Mb1921

CAAX protease self-immunity

Predicted acyl-CoA transferases/
carnitine dehydratase
NADH(P)-binding
NADH(P)-binding
Nucleoside-diphosphate-sugar epimerases
DNA glycosylase
NO
endonuclease III
Deazaflavin-dependent nitroreductase NO
NO
Pyridoxamine 5'-phosphate
Pyridoxamine 5'-phosphate oxidase PPOX class probable F420oxidase
dependent enzyme
Amidohydrolase 2
Amidohydrolase
Predicted metal-dependent hydrolase
of the TIM-barrel fold
Polyketide cyclase/dehydrase
Polyketide cyclase /
START/RHO_alpha_C/PITP/
dehydrase and lipid
Bet_v1/CoxG/CalC (SRPBCC)
ligand-binding domain superfamily
Leucine carboxyl methyltransferase Leucine carboxyl methyltransferase
Leucine carboxyl methyltransferase

Mb1922

Leucine carboxyl methyltransferase Leucine carboxyl methyltransferase

Leucine carboxyl methyltransferase

Mb1927
Mb1931

Aldolase-type TIM barrel

Nitronate monooxygenase
Leucine carboxyl methyltransferase Leucine carboxyl methyltransferase

2-Nitropropane dioxygenase (NPD)

methyltransferase, TIGR00027 family

Mb1915

100

PPE family
NO

Bifunctional nuclease
CocE/NonD hydrolase

Mb1911

HTH-type transcriptional
regulator yodB -like domain

PPE family
NO

Mb1860
Mb1866

Mb1899
Mb1901
Mb1902
Mb1906

CATH

Arsenical Resistance Operon Repressor

similar prokaryotic, metal regulated
homodimeric ..
TIGR03086 family protein

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50

25

75
100
75
50
25
100

NO

75

NO

75

Putative S-adenosyl-L
-methionine-dependent -like domain
Putative S-adenosyl-Lmethionine-dependent -like domain
NO
Putative S-adenosyl-L-

100
100
75
100

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P S Kewate et al. / Journal of Pharmacy Research 2015,9(3),182-189

KEGG NO Interproscan5

PFAM

Mb1948

Beta-lactamase-like

Mb1954

Polyketide cyclase/dehydrase

Mb1955
Mb1965

Phospholipid/glycerol acyltransferase, Acyltransferase

predicted
Class I glutamine amidotransferase-like Putative amidotransferase

Mb1980
Mb1988

HNH nucleases
PIN domain-like

HNH nucleases
NO

Mb1997
Mb2000

VapC family
S-adenosyl-L-methioninedependent methyltransferase-like
VapC family

PIN domain
Methyltransferase domain

Mb2021

Pyruvate/Phosphoenolpyruvate
kinase-like domain

Phosphoenolpyruvate
phosphomutase

Mb2022
Mb2024

Amino acid/polyamine transporter I

Acyl-ACP thioesterase
[Lipid metabolism]
S-adenosylmethionine-dependent
methyltransferases
Predicted kinase
P-loop containing nucleoside
triphosphate hydrolase
VapC family
HNH nucleases
NO
N6 adenine-specific DNA
methyltransferase, N12 class
Erythromycin esterase

Mb2004

Mb2026
Mb2027
Mb2031
Mb2033
Mb2038
Mb2043
Mb2049
Mb2055

Metallo-beta-lactamase
superfamily
Polyketide cyclase / dehydrase
and lipid

PIN domain

Acyl-ACP thioesterase
[Lipid metabolism]
Methyltransferase domain
AAA domain
AAA domain
PIN domain
HNH nucleases
NO
Restriction endonuclease
Erythromycin esterase

CDDBLAST

CATH

Zn-dependent hydrolases,
including glyoxylases
Pyrabactin resistance 1 (PYR1), PYR1like (PYL), regulatory component
of abscisic acid ...
Lysophospholipid Acyltransferases (LPLATs)
of Glycerophospholipid Biosynthesis
Type 1 glutamine amidotransferase
(GATase1)-like domain
HNH nucleases
Predicted nucleic acid-binding protein,
contains PIN domain
PIN (PilT N terminus) domain
S-adenosyl-L-methionine-dependent
methyltransferase-like
PIN domain of the VapC-like UPF0110
protein Mb0640 and homologs
Members of the ICL/PEPM enzyme family.
catalyze either P-C or C-C
bond formation/cleavage.
Amino acid transporters
Acyl-ACP thioesterase
[Lipid metabolism]
S-adenosylmethionine-dependent
methyltransferases
Predicted kinase
Predicted ATPase (AAA+ superfamily)
PIN domain of the Rv0301 toxin
HNH nucleases
Predicted helicase
DEAD-like helicases superfamily
Erythromycin esterase

CONCLUSION
The Bioinformatics based study of protein data available for the M.
bovis has given the opportunity to elucidate enzyme functions in the
hypothetical proteins for linking them to particular metabolic pathway. Study aimed at enzyme function linking in the 468 hypothetical
proteins screened of M. bovis out of those, 347 hypothetical proteins
computationally predicted to harbor enzymatic domains. In those,
254 proteins having confidence limit not less than 50% which represented some better existence probability of them. The functionality
finding in all available hypothetical proteins of M. bovis will remain
the first priority in the future due to its pathogenicity.6 Similar studies
with different bacteria successfully reported the probability of enzyme coding ability of hypothetical proteins in Shigella flexneri,
Bacillus anthracis, H. influenzae and Helicobacter pylori.14 -17 Study
concluded that, available domain search tools linked with rich protein
databases information, proved to be beneficial and filtered out some
of the important enzyme coding proteins out of hypothetical pool of
M. bovis and in future it may assist in relational linking of todays
hypothetical protein in tomorrows functional proteins probably by
involving gene cloning, microarray and marker based experiments.
REFERENCES
1. Lagrange PH, Hurtrel B, and Stach JL, Vaccines against

2.

3.

4.

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6.

7.

8.

methionine-dependent -like domain

Hydroxyacylglutathione
hydrolase -like domain
Abscisic acid receptor
PYL12 -like domain
1-acyl-sn-glycerol-3-phosphate
acyltransferase -like domain
Es1 protein -like domain
NO
NO

%
100
100
100
75
75
50

NO
High-molecular-weight
protein 2 -like domain
NO

75
100

Isocitrate lyase -like domain

100

NO
Ribosomal RNA-processing
protein 8 -like domain
NO
Origin recognition complex
protein 5 -like domain
SMG6 protein -like domain
NO
NO
NO
NO

75

50
75
100
75
100
100
75
25
75
75

mycobacteria and other intracellular multiplying bacteria,

Annales de lInstitut Pasteur DImmunology, 136(2), 1985,
151162.
Buddle BM, Parlane NA, Keen DL et al., Differentiation between Mycobacterium bovis BCG-vaccinated and M.
bovisinfected cattle by using recombinant mycobacterial
antigens, Clinical and Diagnostic Laboratory Immunology,
6 (1), 1999, 15.
Griffin JFT, Chinn DN, Rodgers CR, and Mackintosh CG,
Optimalmodels to evaluate the protective efficacy of tuberculosis vaccines, Tuberculosis, 81(1-2), 2001, 133 139.
World Health Organization, Tuberculosis, WHO, Geneva,
Switzerland, 2014, http://www.who.int/topics/tuberculosis/
en/.
WHO, WHO Global Tuberculosis Report 2013, WHO, 2014,
http://www.who.int/tb/publications/global report/en/.
Muller B, Durr S, Alonso S et al., Zoonotic Mycobacterium bovis-induced tuberculosis in humans, Emerging Infectious Diseases, 19 (6), 2013, 899908.
Garnier T, et al., The complete genome sequence of Mycobacterium bovis, Proc Natl Acad Sci U S A, 100, 2003, 787782.
Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z,
Miller W, Lipman DJ, Gapped BLAST and PSI-BLAST: a
new generation of protein database search programs. Nucleic
Acids Res, 25 (17), 1997, 3389-402.

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Schaffer AA, Aravind L, Madden TL, Shavirin S, Spouge JL,

Wolf YI, Koonin EV, Altschul SF, Improving the accuracy of
PSI-BLAST protein database searches with compositionbased statistics and other refinements, Nucleic Acids Res,
29(14), 2001, 2994-3005.
Marchler-Bauer A et al., CDD: a conserved domain database for interactive domain family analysis, Nucleic Acids
Res, 35(D), 2007, 237-240.
Zdobnov EM, Rolf A, Interproscan- an integration platform
for the signatures recognition methods in InterPro,
Bioinformatics, 17, 2001, 847-848.
Bateman A, et al., The Pfam families database, Nucleic
Acids Res, 30, 2000, 276-80.
Sillitoe I et al., New functional families (FunFams) in CATH
to improve the mapping of conserved functional sites to 3D

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structures, Nucleic Acids Res., 41(Database issue), 2013,

D490-8.
Gore D, In silico prediction of structure and enzymatic
activity for hypothetical proteins of Shigella flexneri,
Biofrontiers, 1 (2), 2009, 1-10.
Gore D, Raut A, Computational function and structural annotations for hypothetical proteins of Bacillus anthracis,
Biofrontiers, 1 (1), 2009, 27-36.
Dogra P, Gore D, Prediction of Enzymatic Function and Structure of H. influenzae Hypothetical Proteins - An In silico
Approach. International Journal of Soft Computing and
Bioinformatics, 1(2), 2010, 67-77.
Gore D, Denge P, Amrute M, Homology Modeling and Enzyme Function Prediction in the Hypothetical Proteins of
Helicobacter pylori - an Insilico Approach, Biomirror, 1-5,
2010, bm-1111251610.

Source of support: Nil, Conflict of interest: None Declared

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