Introduction To Bioorganic Chemistry and Chemical Biology Ch9 Answers

Introduction to Bioorganic Chemistry and Chemical Biology 1
Answers to Chapter 9
(in-text & asterisked problems)
Answer 9.1
Names and acronyms from August 2011 issues of chemistry/biology journals are listed
below. The important point is that most of the papers in the current research literature
involve mysterious acronyms. The goal of this chapter is to familiarize you with those
acronyms that relate to signal transduction pathways that affect gene transcription in
humans.
Journal of Biological Chemistry 286, Issue 34 (2011)
Smad2/3 (a hybrid of C. elegans Sma (small) gene and Drosophila Mad (Mothers against
dpp) gene)
Hepatocyte Nuclear Factor 4
RNA
Photosystem II (PSII)Psb27 (photosystem protein b27) protein complex
ATP synthase
dentin matrix protein
integrin
MSH2 (DNA repair proteins homologous to yeast mutS/hexA)
Cdt2 (protein target of target of Cdc10 transcriptional regulation)
P21Waf1/Cip1 (kinase inhibitor protein 21 encoded by the Waf1/Cip1 genes)
VAMP8 (Vesicle-associated Membrane Protein 8)
SNARE (Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor)
GSK3 (Glycogen Synthase Kinase-3)
PTTG1/Human Securin (protein from Pituitary Tumor-Transforming Gene 1)
Myocyte Enhancer Factor 2C, an Osteoblast Transcription Factor
aurora-A
augmin
Hice1 Protein (Hec1-interacting and centrosome-associated 1)
microtubule
ARD1 (Arabidopsis Acireductone Dioxygenase 1)
Heterotrimeric G protein subunit
Hsp90 (Heat-shock protein 90)
Immunophilin FKBP51 (FK-506 binding protein 51)
PSKs/TAOKs (Prostate-derived Sterile 20-like Kinases)
glucocorticoid
Dig2/RTP801/REDD1 (dexamethasone-induced Gene 2 Protein)
NKCC2 (bumetanide-sensitive Na+,K+,2Cl Cotransporter)
TammHorsfall protein
CaV1.3 channels (voltage-gated calcium channel 1.3)
c-Jun kinase (the Jun oncogene from Avian sarcoma virus was named after Ju nana, the
Japanese word for 17)
Runx2 protein (Runt-related transcription factor 2)
Escherichia coli DinD protein (DNA damage inducible protein D)
RecA (DNA recombinase A)
VAP-1 (Vascular Adhesion Protein-1)
oxime
acetylcholinesterase
2 Introduction to Bioorganic Chemistry and Chemical Biology: Answers to Chapter 9
cytochrome P450SP (a heme enzyme that absorbs light at 450nm)

fatty acid
autotaxin
Egr-1 protein (Early Growth Response 1)
HNF4 (Hepatocyte Nuclear Factor 4)
SHP (Small Heterodimer Partner)
Mef2A transcription factor (myocyte enhancer factor 2A)
GRTH/DDX25 (Gonadotropin-regulated Testicular RNA Helicase)
StAR (Steroidogenic Acute Regulatory protein)
androgen receptor
TGF-1 (Transforming Growth Factor-1)
microRNA 143/145
TLR4 protein (Toll-like receptor 4)
MMP-1 (Matrix Metalloproteinase-1)
Chemical Biology 50, Issue 8 (2011)

retinal
mitochondrial Lon protease
docataxel tubulin
Hsp90 (heat shock protein 90)
JNK (Jun kinase)
Sab kinase (SH3 domain-binding protein that preferentially associates with Btk)
mycobacterial arabinosyltransferase
decaprenylphosphorylarabinose analogues
Kalata B1
Kir2 inward rectifier potassium channels (inwardly rectifying potassium channel 2)
RNA
Biochemistry 50, Issue 33 (2011)

p53 (mammalian tumor suppressor protein (53 kDa))
group 3 LEA protein (Late Embryogenesis Abundant)
nucleoplasmin
histone
prion
P. aeruginosa type III secretion translocators
P. aeruginosa heteromeric transmembrane pores
tyrosyl tRNA synthetase
nitric oxide synthase
Hsp70 (heat shock protein 70)
thioredoxin
rhodopsin
phoborhodopsin
prolylacyl carrier protein oxidase
P. aeruginosa isochorismatepyruvate lyase
GTPase
chloroplast signal recognition particle
S100A4 (calcium-binding proteins soluble in 100% saturated ammonium sulfate)
myosin-IIA
mouse olfactory receptor
glmS riboswitch (glucosamine-6-phosphate activated ribozyme)
glucosamine 6-phosphate -anomer
DNA polymerase
RNA
frataxin
cyclotide
GluA2 AMPA receptor (A2 ion channel receptor that binds to glutamate)
Introduction to Bioorganic Chemistry and Chemical Biology: Answers to Chapter 9 3
Science 333, August 11 (2011)

Fc receptor (receptor that binds to immunoglobulin constant fragment )
agonistic CD40 antibodies (complementarity determining/determinant 40)
Nature 476, Number 7360 (2011)

Lgr5 homologues (leucine-rich repeat-containing G protein-coupled receptor 5)
Wnt receptors (Wnt is a hybrid of the Drosophila Wingless gene and the mammalian
integration 1 gene)
R-spondin
histone
14-3-3 proteins
rice Hd3a florigen (heading date 3a)
MCU (mitochondrial calcium uniporter)
ribosome
Answer 9.2
Cycloheximide would have no effect since mucin release does not involve protein syn-
thesis.
Physostigmine would enhance the effect of acetylcholine by preventing hydrolysis by
acetylcholinesterase.
Atropine would inhibit the rapid release of mucin by antagonizing the effect of acetyl-
choline.
Answer 9.3
Nuclear receptors and gas receptors.
Answer 9.4
The androgen receptor (AR) binds duplex DNA with the sequence AGGTCA; the estrogen
receptor (ER) binds to duplex DNA with the sequence TGTTCT. By connecting the DNA-
binding domain of the androgen receptor to FKBP12 and the DNA-binding domain of
the estrogen receptor to mTOR, the two domains will dimerize and bind to DNA in the
presence of rapamycin. Semiflexible peptide linkers would be needed to connect the
domains; the Ctermini of FKBP12 and mTOR are closer together than the Ntermini.
5'-AGGTCANNNTGTTCT-3' 5'-AGGTCANNNTGTTCT-3'
3'-TCCAGTNNNACAAGA-5' 3'-TCCAGTNNNACAAGA-5'
AR
rapamycin
DBD ER AR ER
DBD DBD DBD
FKBP12
mTOR FKBP12 mTOR
Introduction to Bioorganic Chemistry and Chemical Biology | A9145

Van Vranken & Weiss | 978-0-8153-4214-4
Answer
9.5
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The calculations suggest that roughly more than 1% of enone steroids exist as thiol
adducts, but Nature is cautious. The equilibrium constant for addition to steroid
enones such as testosterone is actually 100-fold less favorable (Keq=0.02) than it is for
3-methylcyclohexenone.
1. How much adduct, x, is present at equilibrium?

K eq
thiol
+ HS adduct
R'
O R O R
S
R'
[A] [R'SH] [AR'SH]
If thiol is depleted: 10-6 M - x 0.005 M - x x
If thiol is not depleted: 10-6 M - x 0.005 M x
2. Set up equations depending on your assumptions

If thiol is not depleted: If thiol is depleted:
[AR'SH] (x) [AR'SH] (x)
K eq = = K eq = =
[A] [R'SH] (10 -6 M - x)(0.005 M) [A] [R'SH] (10 -6 M - x)(0.005 M - x)
x
(10 -6 M - x)(0.005 M) = ... you end up with a quadratic equation; but since
K eq
[R'SH] >> [A], the answers you calculate, will not be
1 very different from the thiol depletion scenario
5x10-9 M2 - (0.005 M) x = x
K eq
5x10-9 M2 1
x= 5x10-9 M2 - (0.005 M + 10-6 M + )x + x2 = 0
1 K eq
(0.005 M + )
K eq
3. Plug in K eq
major
prednisone O progesterone O
adduct adduct
R'
S
O O
less reactive S
R'
K eq ~ 7,000 M-1 K eq ~ 2
x = 0.0097x10-6 M thiol adduct x = 0.0099x10-6 M thiol adduct

0.990x10-6 M prednisone left over 0.990x10-6 M progesterone left over
99% of progesterone left over 99% of progesterone left over
Answer 9.6
H435F,
Van H435K,
Vranken & WeissH435W, and H435R are too big and or
| 978-0-8153-4214-4 too polar to form complementary interactions
with the bulky ethoxy group of QH2. H435G is probably too small and would leave empty space. The
H435A mutant is just right to complement the thyroid mimic QH2.
Answer 9.7
A IL-3, GM-CSF, IL-6, and IL-5
B IL-3, IL-7, IL-6, and IL-5
Answer 9.8
Most noncovalent interactions between small molecules and proteins have fast on-rates and reach
equilibrium very quickly. The fact that inhibition increases with longer incubation times and that the
sterically similar analog is inactive suggests a covalent reaction between stattic and STAT3, which has
numerous free cysteine residues. Cys712 (PDB 1BG1) makes contact with the active site and could
form an adduct with the vinylsulfone moiety.
Cys
Cys
-
S
.. S
O 2N S O 2N S
O O O O
Answer 9.9
The extracellular environment favors disulfide formation. A cysteine residue stabilizes
the dimeric form of the receptor by forming a cystine disulfide crosslink.
R248C or
S249C FGFR3
monomers dimer
D1 D1 D1 D1
D2 D2 D2 D2
C C C C
D3 D3 D3 D3
disulfide
bond
P P

Answer 9.10
At a PD173074 concentration of 1000nM, FGFR1, FGFR2, FGFR3, and VEGFR2 would be
more than 90% inhibited.
Answer 9.11
A FGFR, Shc1, MEK, ERK, and the transcription factor
B FGFR3 and Shc1
C FGFR3, Shc1, and ERK (which is activated through the phosphorylation of both thre-
onine and tyrosine in its activation loop)
Answer 9.12
H H H
N His N His N His
His His His
H N N+ H N N+ H N N+
N: N+ N+
H H - H H H
H
DAG DAG .. DAG
HO O O HO O: O HO
O O
O- O- O-
P P -HO PO P
-HO
3PO O H H -HO
3PO O H H 3 H H
O +N O +N O -O +N
-HO H Arg -HO H Arg -HO PO H Arg
3PO OH N N 3PO OH N N 3
OH N N
H H H
salt bridge H H H
H H H
N His N His N His
His His His
H N N H N ..N H N N
N+ N+ N+ +
H H H H
H H H H
+ DAG O - ..O
HO O HO HO
O - O - O -
O O O
-HO PO P - HO3PO P -HO PO P
3 H H H H 3 H H
O -O +N O O +N O O +N
-HO PO H Arg -HO H Arg -HO PO H Arg
3 N N 3PO N N 3 N N
OH OH OH
H H H
H H H
H H H
N His N His N His
His His His
H N N+ H N N+ H N N+
N: N+ N+
H H - H H H
H ..
HO O H HO O H HO H
O O OO
O- O- O-
P P -HO
3PO
P
-HO
3PO O H H - HO3PO O H H .. H H
O +N O +N O -O +N
H Arg H Arg -HO H Arg
-HO
3PO OH N N -HO
3PO OH N N 3PO N N
OH
H H H
H H H
Answer 9.13
O O O O O
H H H H H
O O O O O
AcO AcO AcO AcO AcO
MeO MeO MeO MeO + MeO
O O O OH OH
H : A-
O O .. O O O
+ +
H A H H2N H N R H :N R
O O R O H O H O N
H : A- H + R
an equally acceptable mechanism would involve... or...

O O O O
H H H A +
etc. H H etc.
O O O: OH
AcO AcO AcO AcO
MeO H A MeO H MeO MeO
O O O O
O O :Nu O O
+ + :Nu
O O O O
Answer
9.14
AlF3 can form a non-hydrolyzable adduct with GDP that mimics GTP.
O
O
F - O O O- N
F N F O NH
- OO O - NH Al
O P P
Al P P F - O O O N N NH2
:O O O N N NH2 O
FF O
HO
HO OH
OH non-hydrolyzable
Answer 9.15
www.garlandscience.com
Because design byadenylyl
forskolin activates cyclase and a CB1 agonist reverses the effect of forskolin, it suggests that CB1 is coupled to Gi,
www.blink.biz
an inhibitor of adenylyl cyclase. If the CB1 receptor is coupled to Gi, the effect of 9-tetrahydrocannabinol would be inhibited by
pertussis toxin.
Answer 9.16
Various authors have suggested that aldehydes bind to the I7 odorant receptor as an
iminium ion (also known as a Schiff base), but there is no mechanistic evidence for or
against the hypothesis.
helix
4 K164
Y107
HO helix
3
+HN
O
O HO
D204
-
Y264
helix helix
5 6
F205
F262
Answer 9.17
+
+
- - -
+ : : :
+
+
+
- .. - .. - ..
Van Vranken9.18
Answer & Weiss | 978-0-8153-4214-4
(From J.T. Gerig et al., J. Inorg. Biochem. 31:113121, 1987. With permission from Elsevier.)
CO2-
N
O
OH OH
O
O O
N Ca N
O O
O O
O O
Answer 9.19
This
is similar to the photodeprotection
www.garlandscience.com design by www.blink.biz of oligonucleotides in Chapter 3.
H -A: H
O O- O O- O O- O-
H N
+
H N
+ -O2C .. N+ +O N
-O -O -O2C
2C h 2C RO RO
RO RO
-
O: H A OH OH + OH OH
O N O N :O N O N O N
-O2C -O2C -O2C -O2C -O2C
RO O O .. HO
H2O
R' O
.. R' O+
H A H
-A: H -
O O- O O- O-
-O2C N
+ -O2C .. N+ O N
O: H A OH OH + OH
O N O N :O N O N O N
-O2C -O2C -O2C -O2C -O2C
RO O O .. HO
H2O
8 Introduction to Bioorganic Chemistry and Chemical Biology:
R' Answers
O to Chapter
R' O +9 ..
H A H
-A: H -
O O- O O- O-
-O2C N
+ -O2C .. N+ O N
or... -O2C
RO RO RO

Answer
Van Vranken &9.20
Weiss | 978-0-8153-4214-4
The product is undoubtedly
www.garlandscience.com a conjugate addition product.
design by www.blink.biz The stereochemistry at the
newly formed stereogenic center to the carbonyl cannot be readily predicted without
knowledge of the interaction between kamebakaurin and the binding site.
OH O OH O
-
H S
:S
HO Cys HO HO Cys
HO
OH OH
Van Vranken &
*Answer Weiss | 978-0-8153-4214-4
9.23
A Tyrphostin would inhibit all elements of these pathways.
B Wortmannin inhibits PI3K, the first enzyme in the apoptotic pathway.
C Rapamycin would selectively inhibit TOR and the effects on protein synthesis.
*Answer 9.25
R R R R
H h h
H H H >100C H H H H
HO HO HO
photo-
HO pyrocalciferol pyrocalciferol precalciferol calciferol
*Answer 9.27
The residues that contribute most significantly to binding are positively charged lysines
and arginines. It is therefore probably interacting with negatively charged aspartate or
glutamate residues on the receptor.
*Answer 9.29
There is some ambiguity in the P+3 position, but on the basis of the intensity of the
spots, the best consensus sequence would be
*
XXPXYVNIR
The authors were cautious in their interpretation of preferences and counted only spots
that bound strongly to Grb2.
* I
XXXXYI V NLX
V
*Answer 9.31
The simple way to do this is to load a suitably protected lysine onto an acid-labile resin
and then deprotect both the -amino group and the side chain. Then one can perform
traditional solid-phase peptide synthesis using twice the normal amount of Fmoc-
protected amino acids. Finally the peptide can be cleaved under mild acid conditions.
(Arg protected by Pbf or Pmc) acid cleavage: e.g.,
1:1
O (Arg protected O
by Pbf or Pmc) acid cleavage:
CF 3CO2H/ e.g.,
SPPS to Bioorganic(Arg
Introduction H
Chemistry and or Pmc) Biology:acid
Chemical cleavage:
Answers e.g., 9
2Cl2 to Chapter 9
protected
N by Pbf CH 1:1
H2N
O O H2N-VPPPVPPRRR O O CF 3CO 1:1H/
H 2
NH2 O SPPS NH O CF 3Cl
CH CO2H/
H2N SPPS N H 2 2
H2N here
grow O X H
H2N-VPPPVPPRRR2 N -VPPPVPPRRR
N Oacid- X CH 2Cl2
O H2N-VPPPVPPRRR O
(Arg protected by Pbf or Pmc) labile acid cleavage: e.g.,
NH2 NH
NH X H2N-VPPPVPPRRR NH acid- X 1:1
grow here 2
X H2N-VPPPVPPRRR X 3CO2H/
grow here O O acid- CF
SPPS H labile
H2N N labile CH2Cl2
O H2N-VPPPVPPRRR O
NH2 NH
grow here X H2N-VPPPVPPRRR acid- X
labile
The key challenge is how to deprotect the N and N amino groups without cleaving the amino acid from the resin. The authors
attached N-Fmoc-N-Fmoc lysine to a commercial Wang resin through a carbodiimide coupling and then removed the Fmoc
O Wang linker
groups using piperidine/DMF. HMB resin O
FmocHN H2N
O O Wang linker HMB
H resin O O H SPPS
O Wang linker
FmocHN NH NHMB resin piperidine H2N NH2 O N
FmocHN Fmoc Oacid- O H2N O O
SPPS
O H O H
NH labile O NH DMF
piperidine NH2 O NH SPPS
Fmoc NH acid- O N piperidine NH2 O N
Fmoc
O Wangacid-
linker OHMB resin O O
labile O DMF O
FmocHN labile O DMFH2N O
O O SPPS
H H
NH N piperidine NH2 N
Fmoc acid- O O
labile O DMF O
Alternatively one could attach the first amino acid to acid-labile chlorotrityl resin and follow the same sequence of steps as with
the Wang resin.
Grb2 peptidimer ligand

H
Grb2 peptidimer
N ligand CO2-
Grb2 peptidimer ligand
H2N-VPPPVPPRRR
C-SH3 N-SH3 H
NH NHCO2-
H2N-VPPPVPPRRR
H2N-VPPPVPPRRR N CO2-
C-SH3 N-SH3 H2N-VPPPVPPRRR
C-SH3 N-SH3 Grb2 peptidimer ligand NH
H2N-VPPPVPPRRR NH
H2N-VPPPVPPRRR
H
N CO2-
H2N-VPPPVPPRRR
C-SH3 N-SH3 NH
H2N-VPPPVPPRRR
The least desirable solution (that would still work) would be to attach the very common N-Fmoc-N-Boc lysine to an acid-stable
resin such as Merrifield resin and then use TFA to cleave the Boc side chain without cleaving the benzyl ester linkage.
very acid-labile
O Merrifield resin O
H i) CF3 CO2 H anhydrous
veryOacid-labile
N H2N SPPS
very acid-labile O OMerrifield resin (remove Boc) O O
HF
H O Merrifield resin i) CF3 CO2 H anhydrous
O O NH NH H2N NH2 O SPPS
O N Fmoc O not ii) i)piperidine
(remove CF3 CO 2H
Boc) H2N O SPPS
anhydrous
HF
O
very DMF (remove Boc) O HF
O
very acid-labile NH NH2
O Fmoc NHacid-labile
not (remove Fmoc)
ii) piperidine NH2
O Merrifield
Fmoc notresin DMF ii) piperidine O
H very i) CF
very 3 CO
DMF
(remove 2 H Fmoc) anhydrous
O N acid-labile (remove H2N
Boc) Fmoc)
SPPS
HF
O acid-labile (remove O
Introduction
O to BioorganicNH Chemistry and Chemical Biology | A9162 NH2
Van Vranken & WeissFmoc
| 978-0-8153-4214-4
not ii) piperidine
Introduction to Bioorganic design
www.garlandscience.com byvery
Chemistry www.blink.biz DMF| A9162
and Chemical Biology
VanIntroduction to Bioorganic
Vranken & Weiss Chemistry and Chemical Biology
acid-labile
| 978-0-8153-4214-4 (remove | A9162
Fmoc)

*Answer 9.33
high energy excited state
OMe OMe * OMe
OMe
NO2 NO2 NO2 B:
NO2
.. H
N h +N +N H N
N .. N+ N+
OO O- O O- O - O O- N+
:O- O O O-
CO2- CO2- CO2- +H3N
+H
+H
3N
+H
3N 3N -O
2C
aromatic OMe OMe

H H
OMe OMe
NO2 NO2 B: NO2 NO2
N ..N N+ N
H H
N .. N N N
O O .. O - B H .. O O O
+H3N
O O-
-O +H3N
2C free
-O
2C glutamate
*Answer
9.35
JAK O
JAK O O-
SH O O O
OH
S OH
OH
S
O SH
JAK
JAK
*Answer
9.37
NH2
O O
-O P O- -O P O-
:S
O O
O O O
H .. H H
N N N
R N R' R N - R' R N R'
H H H H
O O O
..
B- NH2 NH2 NH2
O
S
O
S ~
~ O
- H H H
N N N
R N .. R' R N R' R N R'
H H H
O O O
H
B lysine
*Answer
9.39
Me Me OH
Me Me
liver enzyme; OH
synthetic kidney enzyme
vitamin D2 ligand for the
(ergocalciferol) vitamin D receptor
HO HO

Introduction To Bioorganic Chemistry and Chemical Biology Ch9 Answers

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Introduction To Bioorganic Chemistry and Chemical Biology Ch9 Answers

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Introduction to Bioorganic Chemistry and Chemical Biology 1

cytochrome P450SP (a heme enzyme that absorbs light at 450nm)

Chemical Biology 50, Issue 8 (2011)

Biochemistry 50, Issue 33 (2011)

Science 333, August 11 (2011)

Nature 476, Number 7360 (2011)

Introduction to Bioorganic Chemistry and Chemical Biology | A9145

1. How much adduct, x, is present at equilibrium?

2. Set up equations depending on your assumptions

x = 0.0097x10-6 M thiol adduct x = 0.0099x10-6 M thiol adduct

Introduction to Bioorganic Chemistry and Chemical Biology | A9148

an equally acceptable mechanism would involve... or...

Introduction to Bioorganic Chemistry and Chemical Biology | A9155

Grb2 peptidimer ligand

Introduction to Bioorganic Chemistry and Chemical Biology | A9162

aromatic OMe OMe

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