UNIVERSITY OF THE PHILIPPINES MANILA

COLLEGE OF ARTS AND SCIENCES

DEPARTMENT OF BIOLOGY

A not every micronutrient listed is required by all cells some metals

listed are found in enzymes or cofactors present in only specific
microorganisms
B needed in greater amounts than other trace metals

BIOLOGY 120 (General Microbiology)

Lecture 2.1 Microbial Nutrition and Metabolism
By:
Lani Manahan-Suyom, MSc
SS AY 2013-2014
Outline
1.0 Microbial Nutrition
1.1 Nutrition and Cell Chemistry
1.2 Nutrient Uptake
1.3 Culture Media
2.0 Metabolism
Microorganisms require about ten elements in large quantities, because
they are used to construct carbohydrates, lipids, proteins, and nucleic acids.
Several other elements are needed in very small amounts and are parts of
enzymes and cofactors
Biochemical components of cells

Water: 80% of wet weight

Dry weight
o Protein
40-70%
o Nucleic Acid 13-34%
o Lipid
9-15%
o Also monomers, intermediates and inorganic ions
Macronutrients

Cells make proteins, nucleic acids and lipids

Macronutrients
o Macromolecules, metabolism
o CHON SP K Mg Fe
o Sources

Organic compounds

Inorganic salts

Micronutrients

Elements needed in trace quantities

o Co, Cu, Mn, Zn, V
o Enzymes
o Tap water

Most of them are part of enzymes

o Iron cytochrome, carboxylases
Micronurients (trace elements) needed by microorganisms
Boron (B)
Autoinducer for quorum sensing in bacteria also
found in some polketide antibiotics (creates biofilm)
Chromium
Possible but not proven component for glucose
(Cr)
metabolism (necessary in mammals)
Cobalt (Co)
Vitamin B12; transcarboxulase (only in propionic
acid bacteria)
Copper (Cu)
In respiration, cytochrome oxidase; in
photosynthesis, plastocyanin, some superoxide
dismutases
b
Iron (Fe)
Cytochromes; catalases; peroxidises; iron-sulfur
proteins; oxygenases; all nitrogenises
Manganese
Activator of many enzymes; component of certain
(Mn)
superoxide dismutases and of the water splitting
enzyme in oxygenis phototrophs (photosystem II)
Molybdenum Certain flavin-containing enzymes; some
(Mo)
nitrogenises, nitrate reductases, sulphite oxidases,
DMSO-TMAO reductases; some formate
dehydrogenases
Nickel (Ni)
Most hydrogenases; coenzymes F430 of
methanogens; carbon monoxide dehydrogenases;
urease
Selenium
Dormate dehydrogenase; some hydrogenases; the
(Se)
amino acid selenocysteine
Tungsten
Some formate dehydrogenases; oxotransferases of
(W)
hyperthermophiles
Vanadium(V) Vanadium nitrogenises; bromoperoxidase
Zinc (Zn)
Carbonic anhydrase; alcohol dehydrogenase; RNA
and DNA polymerases and many DNA binding
proteins

Growth Factors

Biotin Carboxylation (Leuconostoc)

Cyanocobalamin or Vit B12 Molecular rearrangements (Euglena)

Folic Acid one carbon metabolism (Enterococcus)

Pantothenic Acid fatty acid metabolism (Proteus)

Pyridoxine or Vit B6 transamination (Lactobacillus)

Niacin precursor of NAD and NADP (Brucella)

Riboflavin or Vit B2 precursor of FAD and FMN (Caulobacter)

Thiamine or Vit B1 aldehyde group transfer (Bacillus anthracis)

Groups of microorganisms: nutritional requirement
Energy Sources
Phototrophs
Light
Chemotrophs
Oxidation of organic/inorganic
compounds
Hydrogen and Electron Sources
Lithotrophs
Reduced inorganic molecules
Organotrophs
Organic molecules
Carbon Sources
Autotrophs
CO2 sole or principal
biosynthetic carbon source
Heterotrophs
Reduced, preformed, organic
molecules from other organisms
Major Nutritional
Types
Photolithotrophic
Autotrophy

Photoorganotrophic
Heterotrophy

Sources of Energy,
Hydrogen/Electrons
and Carbon
Light energy
Inorganic
hydrogen/elect
ron donor
CO2 carbon
source

Representative
Microorganisms

Chemolithotrophic
Autotrophy

Chemoorganotrophic
Heterotrophy

Light energy
Organic
hydrogen/elect
ron donor
Organic carbon
source (CO2
may also be
used)
Chemical
energy source
(inorganic)
Inorganic
hydrogen/elect
ron donor
CO2 carbon
source
Chemical
energy source
(organic)
Organic
hydrogen/elect
ron donor
Organic carbon
source

Algae
Purple and
green sulphur
bacteria
Blue-green
algae
(cyanobacteria)
Purple nonsulfur bacteria
Green nonsulfur bacteria

Sulfur oxidizing
bacteria
Hydrogen
bacteria
Nitrifying
bacteria
Iron bacteria
Protozoa
Fungi
Most nonphotosynthetic
bacteria

Passive Diffusion
Passive diffusion is the process in which molecules move from a region
of higher concentration to one of lower concentration as a result of
random thermal agitation. A few substances, such as glycerol, can cross
the plasma membrane by passive diffusion

Chemoorganotrophs are by definition heterotrophs.

Most chemolithotrophs and phototrophs are autotrophs.
Autotrophs are sometimes called primary producers
o they synthesize new organic matter from CO2
Virtually all organic matter on Earth has been synthesized by primary
producers, in particular, the phototrophs

Properties of Microbial Photosynthetic Systems

Property
Cyanobacteria Green and Purple
Bacteria
Photopigment Chlorophyll
Bacteriochlorophyll
O2
Yes
No
Production
Electron
H2O
H2, H2S, S
Donors
Carbon
CO2
CO2
Source
Primary
ATP + NADPH
ATP
Products of
Energy
Conversion
Chemoautotroph
Bacteria
Electron donor
Alcaligens and
Pseudomonas
sp.
Nitrobacter
Nitrosomonas
Desulfovibrio
Thiobacillus
denitrificans
Thiobacillus
ferooxidans

Purple Nonsulfur
Bacteria
Bacteriochlorophyll
No
H2, H2S, S
Organic/CO2
ATP

Production

H2

Electron
acceptor
O2

NO2
NH4
H2
S, H2S

O2
O2
SO4
NO3

NO3, H2O
NO2, H2O
H2O, H2S
SO4, N2

O2

Fe , H2O

2+

Fe

Facilitated Diffusion
The rate of diffusion across selectively permeable membranes is greatly
increased by the use of carrier proteins, sometimes called permeases,
which are embedded in the plasma membrane. Since the diffusion
process is aided by a carrier, it is called facilitated diffusion. The rate of
facilitated diffusion increases with the concentration gradient much
more rapidly and at lower concentrations of the diffusing molecule
than that of passive diffusion
Can be against or with concentration gradient
A Model of Facilitated Diffusion
The membrane carrier can change conformation after binding an
external molecule and subsequently release the molecule on the cell
interior. It then returns to the outward oriented position and is ready
to bind another solute molecule
Because there is no energy input, molecules will continue to enter only
as long as their concentration is greater on the outside
Symport and Antiport

H2O

3+

Nitrifying Bacteria

Uptake of Nutrients
Nutrient molecules frequently cannot cross selectively permeable
plasma membranes through passive diffusion and must be transported
by one of three major mechanisms involving the use of membrane
carrier proteins
1.
2.

Phagocytosis Protozoa
Permeability absorption most microorganisms

Simple transport
o Passive transport or simple diffusion
o Facilitated diffusion*

Group translocation**

ABC transporter**
*could transport agains or with the concentration gradient
**active transports against concentration gradient

Active Transport
Active transport is the transport of solute molecules to higher
concentrations, or against a concentration gradient, with the use of
metabolic energy input

Group Translocation
The best-known group translocation system is the
phosphoenolpyruvate: sugar phosphotransferase system (PTS), which
transports a variety of sugars into prokaryotic cells while
Simultaneously phosphorylating them using phosphoenolpyruvate
(PEP) as the phosphate donor
PEP + sugar (outside) pyruvate + sugar-P (inside)
Mechanism of the phosphotransferase system of E. Coli
o For glucose uptake, the system consists of five proteins:
Enzyme (Enz) I, Enzymes IIa, IIb, and IIc, and HPr. A phosphate
cascade occurs from phosphoenolpyruvate (PE-P) to Enzyme
IIc and the latter actually transports and phosphorylates the
sugar. Proteins HPr and Enz I are nonspecific and transport
any sugar. The Enz II components are specific for each
particular sugar

Major Classes of Media

Defined
o Precise amounts of highly purified inorganic or organic
chemicals
o Exact composition (in both qualitative and quantitative sense)
is known

Complex
o Employ digests of microbial, animal or plant products, such as
casein (milk protein), beef (beef extract), soybeans (tryptic
soy broth), yeast cells (yeast extract), or any of a number of
other highly nutritious yet impure substances

ABC Transporter

Other Classification of Media

According to use
o Enriched
o Selective
o Differential
o General purpose

Fluidity
o Solid
o Semi solid
o Liquid

ABC stands for ATP binding cassette

ABC transporters exist for the uptake of organic compounds (sugars
and amino acids, inorganic nutrients such as sulfate and phosphate,
and trace metals)

Proteins involved
o Periplasmic binding protein high affinity to substrate even
at low concentration (less than 1 micromolar)
o Membrane spanning transporter forms the transport
channel
o ATP hydrolyzing protein supply energy

Has high affinity for to the molecule it needs to transport (even if

molecule level is really low)
Simple Comparison of Transport Systems

Development of Solid Medium

Before agar
o Liquid medium

Potato slices
o Robert Koch (1881) used boiled potato, sliced
o Not all bacteria grew well

Gelatin
o Frederick Loeffler
o Meat extract medium + gelatine
o But gelatine liquid at RT

Agar
o Fannie Hesse (1882)
o Agar used for jams and jelly
o Generally not metabolized by microbes
o Liquefies at 100C, solidifies at 40C
Anaerobic Condition for Growth

Reducing media
o Contain chemicals (thioglycollate of oxyrase) that combine O2
o Heated to drive off or use up O2

Anaerobic Jar
Types of Anaerobic Bacteria

Facultative anaerobes

Obligate anaerobes

Microaerophyllic anaerobes certain amount of oxygen tolerated

Culture Media nutrient solutions used to grow microorganisms

mostly for chemotroph organism
Despite the fact that microbiologists have been growing
microorganisms in laboratory cultures for over 125 years, most
microorganisms in nature have yet to be cultured, leaving many
challenges to the microbiologist today

Anaerobic Chambers/Incubators
Anaerobic Jars
o with anaerobic pack (releases CO2 and H and produces
water)
o gas generators
o candle jar usually more effective in microaerophyllic (flame
uses up all the oxygen and flame dies)

An Overview of Metabolism

Glycolysis (Embden-Meyerhof-Parnas pathway)

Stage 1 preparatory reactions

o These are not redox reactions and do not release energy
o Lead to the production of a key intermediate of the pathway

Stage 2 production of NADH, ATP and Pyruvate

o Redox reactions
o Energy is conserved in the form of ATP, and two molecules of
pyruvate are formed
o Redox balance has not yet been achieved
o Pyruvate formed here

Stage 3 consumption of NADH and production of Fermentation

products
o Redox reactions occur once again
o Fermentation products are formed

Metabolism
total of all chemical reactions occurring in the cell
A simplified view of cell metabolism depicts how catabolic degradative
reactions supply energy needed for cell functions and how anabolic
reactions bring about the synthesis of cell components from nutrients
Note that in anabolism, nutrients from the environment or those
generated from catabolic reactions are converted to cell components,
whereas in catabolism, energy sources from the environment are
converted to waste products
Enzymes

Biological catalyst
o Lowers activation energy of a reaction

Enzymes are generally much larger than the substrates

Present in small amount because they are reusable

Active site the portion of the enzyme to which the substrate binds
Catabolism

Fermentation
o Anaerobic catabolism
o Organic compound is both an electron donor and an electron
acceptor
o ATP is produced by substrate level phosphorylation

Respiration
o Catabolism in which a compound is oxidized
o O2 (or an O2 substitute) as the terminal electron acceptor
o Usually accompanied by ATP production by oxidative
phosphorylation

more ATP is produced in respiration than in fermentation and thus

respiration is the preferred choice. But many microbial habitats lack O2
and in such habitats, fermentation is the only option for energy
conservation by chemoorganotrophs

Kreb Cycle (Citric Acid Cycle)

NADH produces 2-3 ATPs

Respiration and Electron Transport

Respiration in which molecular oxygen or some other oxidant serves as

the terminal electron acceptor

The discussion of respiration deals with both the carbon and electron
transformations
o (1) The biochemical pathways involved in the transformation
of organic carbon to CO2
o (2) The way electrons are transferred from the organic
compound to the terminal electron acceptor, driving ATP
synthesis at the expense of the proton motive force

Electron Transport

Electron transport systems are composed of membrane associated

electron carriers. These systems have two basic functions
o (1) to accept electrons from an electron donor and transfer
them to an electron acceptor
o (2) to conserve some of the energy released during electron
transfer for synthesis of ATP
In bacteria ETC happens in plasma membrane not mitoch. membrane
Types of Oxidation-reduction enzymes involved in electron transport
1. NADH dehydrogenases
2. Riboflavin containing electron carriers generally called flavoproteins
3. Iron sulphur proteins
4. Cytochromes
*in addition, one class of nonprotein electron carriers is known, the lipid
soluble quinines
ATP and Cell Yield

The amount of ATP produced by an organism has a direct effect on cell

yield

Cell yield is directly proportional to the amount of ATP produced

Energy costs for assembly of macromolecules are much the same for all
microorganisms

Production of ATP is specific to a strain which is related to its cell yield

Energy Storage

Most microorganisms produce insoluble polymers that can later be

oxidized for the production of ATP

Importance of polymer formation

o Potential energy is stored in a stable form
o Insoluble polymers have little effect on the internal osmotic
pressure of cells
*Storage polymers make possible the storage of energy in a readily
accessible form that does not interfere with other cellular processes
The Proton Motive Force

Carriers in the Electron Transport Chain (ETC) are

o Arranged in increasing positive reduction potential
o Oriented in such a way that as e- are transported, protons are
separated from eo The final donating the electrons plus protons to a terminal
electron acceptor such as O2
o H+ are extruded to the outer surface of the membrane
o H+ originate from two sources

NADH

Dissociation of water into H and OH2??

o The extrusion of H+ to the environment results in the
accumulation of OH2 on the inside of the membrane
o As a result the two sides of the membrane differ in both
charge and pH
o Formation of an electrochemical potential across the
membrane = PMF
ATP Synthase (ATPase)

The use of PMF to generate ATP

Catalyzed a reversible reaction between ATP and ADP + P

Two components
o F1 complex carries out the chemical functions
o F0 complex ion translocating functions

Composition
o F1 complex - 3 3
o F0 complex - a,b2, c12

F0 generates the torque that is transmitted to F1

Assignment: Read Similarities of the ATP Synthesis via ATPase and

flagellar locomotion

Anabolism: biosynthesis

Sugar

Amino Acids

Fatty Acids

Regulation of the Activity of Enzymes: Feedback Inhibition

Reaction shut off because of an excess of the end product

Allosteric enzymes has 2 binding sites, the active site (where substrate
binds) and the allosteric site, where the end product of the pathway
binds

Allosteric Site

With 2 protein binding sites

Without end product in the environment - active site can accept a

substrate, so chemical production will proceed

If end product is in excess it will bind to the allosteric site and prevent
substrate from binding to active site
Regulation of the Activity of Enzymes: Covalent Modification

A) when cells are grown with excess ammonia, glutamine synthetase is

covalently modified by adenylylation as many as 12 AMP groups can be
added. When cells are ammonia-limited, the groups are removed,
forming ADP

B) Adenylylated GS subunits are catalytically inactive so the overall GS

activity decreases progressively as more subunits are adenylylated.
If end product is present, it will bind to the enzyme but when it binds,
the enzyme activity will just go down
The more product that binds, the less active the enzyme activity will be
Activity of enzyme not totally shut down