Download Endospore production by Bacillus subtilis The Bacterial Endospore

Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
Endospore production by
Bacillus subtilis
Microbial Diversity
Many bacterial species have the capacity to
differentiate: in other words- produce
subpopulations with different shapes and/or
function
• Endospores
– Highly differentiated cells resistant to heat, harsh
chemicals, and radiation
– “Dormant” stage of bacterial life cycle
Sporulation:
Bacillus and Clostridium
– Ideal for dispersal via wind, water, or animal gut
Heterocyst formation: filamentous cyanobacteria
(Anabaena)
– Only present in some gram-positive bacteria
Represents a survival mechanism
Differences between Endospores
and Vegetative Cells
The Bacterial Endospore
Spore formers can be classified by
where in the vegetative cell the spore forms
Figure 4.38
Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
Differences between Endospores
and Vegetative Cells
The Life Cycle of an EndosporeForming Bacterium
“mother cell”
“fore spore”
cell lysis
One of the reasons Bacillus anthracis
is a good potential biowarfare agent
Figure 4.39
Consists of highly cross-linked proteins (kratin-like) Not all spore formers produce one
70 different proteins make up 30% total spore proteinComposed mainly of proteins and
glycoproteins
Endospore Structure
Hydrophobic:
protection
adherence
pathogenesis
– Structurally complex
– Contains dipicolinic acid
– Enriched in Ca2+
– Core contains small-acid soluble proteins (SASP)
Region of loosely cross-linked peptidoglycan
Helps maintain dehydration of core region
Synthesized by mother celldoesn’t require active protein synthesis when assembled.
Primary role is protection from chemical, predators
Figure 4.41
Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
Dipicolinic Acid (DPA)
Stages in B. subtilis endospore formation
Takes about 8h to occur
Vegatative cell
DNA condenses into
the axial filament
10-15% dry weight of a endospore
Beginning of asymmetric
cell division
Small Acid Soluble Proteins (SASPs)
Bind to DNA and protect molecule, and provide a
carbon and energy source for germination
Fig. 4.49
Core also has DNA repair enzymes
The point of no return
Figure 4.43
Several things happening
-engulfment of forespore
-synthesis of cortex region
-exosporium synthesis
Stage 4 involves synthesis
of the coat layer
that surrounds the endospore
Culture starts to get resistant to stress
a)Continued synthesis of cortex
b)Accumulation of DPA/Ca
c)Production of SASPs
Figure 4.43
What these stages look like
under the electron micrograph
VC
II
III
V
Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
What controls initiation of Sporulation?
Lets talk about the regulation of sporulation …
Sporulation in B. subtilis is a cascade
of gene expression
events in both the mother cell and the developing spore
Onset is brought about by starvation (C,P,N)
Important that cell density is high
Elaborate decision making process
P
P
A cascade of sigma factors are
important during spore formation
Spore formation involves signaling between the
fore spore and mother cell compartments.
RNA polymerase: enzyme involved in transcription
β
β‘
2α
sigma
SpoIIAA
core
holoenzyme
σ
Different σ’s direct core RNA pol to bind to a
different set of genes
Protease
SpoIIE
Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
Heterocyst Formation
Nitrate (NO3-) Metabolism
Assimilative Pathway
(plants, fungi,
bacteria)
Only certain prokaryotes have the ability to
carryout nitrogen fixation
NO3Nitrate reductase
NH3 -repressed
Dissimilative pathway
(bacteria only)
NO2(nitrite)
Anoxia-derepressed
NH3
Dissimilative reduction to
ammonia; some bacteria
Nitrite reductase
N2 -------> NH3 (NH4+)
NO3organic nitrogen
[NH2OH]
NO (nitric oxide)
Two groups
Free-living nitrogen fixers
Symbiotic nitrogen fixers
NH3 (ammonia)
R-NH2 (organic N)
Assimilation of Inorganic Nitrogen
N2O (nitrous oxide)
N2
Transamination Reactions
Two Mechanisms:
1. Glutamic dehydrogenase
α-ketoglutarate + NH3 -----> glutamic acid
2. Second mechanism uses a 2 enzyme system: Lglutamine synthetase and glutamate synthetase or
GOGAT enzyme.
GS
Glutamic acid + NH3 + ATP -------> glutamine
GOGAT
Glutamine + α-ketoglutarate --------> 2 glutamic acids
Glutamic acid + OAA
Glutamic acid + pyruvate
Aspartic acid + α-ketogluterate
Alanine + α-ketogluterate
Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
Nitrogen Fixation
Nitrogenase Complex
-Dinitrogenase reductase
-Dinitrogenase
Nitrogenase sensitive
to oxygen
When fixed N becomes limiting …
Heterocyst formation
Nitrogen fixation
Anabaena heterocyst
spaced every 10-20 cells
Microm 410 Fall 2009: Endospores & heterocysts
Dr. Parsek
Oxygenic Photosynthes
O2
glycolipids
Cyclic photophosphorylation
Non-cyclic photophosphorylation
Fig. 17.19
Fig. 12.80
Only a few cells undergo differentiation
Heterocysts can not differentiate back
into an oxygenic photosynthetic cell!
Calvin-Benson Cycle
Ribulose bisphosphate
carboxylase (RubisCO)
microplasmodesmata
phosphoribulokinase
GOGAT
glu
Fig. 17.22
Phototrophic heliobacteria: can carry out anoxygenic photosynthesis, sporulate,
and fix nitrogen!