Transcript INTRODUCTION TO THE STUDY OF MICROBIAL PHYSIOLOGY

CHEMICAL COMPOSITION
OF THE BACTERIAL CELL
Size of the “average cell”
• A given strain (E. coli B/r) in steady-state growth
(unchanging rate and constant average cell size and
composition) under specified conditions (balanced
growth)
• The average cell: a rod-shaped body with diameter
and length of approximately 1 and 2 μm,
respectively
• Total volume of an average cell: approximately 9 x
10-13 mL (0.9 femtoliters). The periplasmic space
forms a full 30% of the cell volume
Meaning of the “average cell”
• Approximately 44% along the cell cycle in age
• Approximately 33% larger than when it was born (If
individual cells increase in mass exponentially)
• The idealized frequency distribution (relative number
of cells) of cell age in a steady-state culture
increasing by binary fission is given by f(x) = 21-x,
where x is the age of cells measured on the scale
from 0 to 1 cell generation time
Weight of the “average cell”
• Total weight of an average cell: 9.5 x 10-13 g
• Total dry weight of an average cell: 2.8 x 10-13 g
• Approximately 70% of the packed cell mass is water
(c.a. 90% in cells of higher organisms)
• 1.05 x 1012 cells / g of wet biomass
• Density of the average cell: 1.06 g/mL
Composition of the “average cell”
• Elemental assay of dry cells: approximately 50%
C, 20% O, 14% N, 8% H, 3% P, 2% K, 1% S, 0.2%
Fe, 0.05% each of Ca, Mg, Cl, and a total of 0.3%
trace elements including Mn, Co, Cu, Zn, Mo
• Components: protein 55%, ribosomal RNA 16.7%,
transfer RNA 3%, messenger RNA 0.8%, DNA 3.1%,
lipids 9.1%, LPS 3.4%, peptidoglycan 2.5%, building
block metabolites and vitamins 2.9%, inorganic ions
1.0%
• It will contain 18,700 ribosomes and a little over 2
million molecules of protein, of which there are
between 1000 and 2000 different varieties
Genome size of the “average cell”
• Chromosomal DNA of E. coli: a ccd molecule of 4,720
kbp / approximately 1 mm long
• Although considered haploid, will contain two copies
of the chromosome when growing rapidly
• Maximal number of protein-encoding genes: cannot
possibly be more than 4,300 protein-encoding gene
(average mw. of proteins is 40,000 / average mw. of
an amino acid residue in protein is 110 / the average
protein has 364 amino acids / the average gene size
is 1.1 kbp)
• The genome size of cells of the genus Mycoplasma is
approximately one-fifth the size of that of E. coli
http://www.sci.sdsu.edu/~smaloy/MicrobialGenetics/topics/chroms-genes-prots/genomes.html
Minimum genome size of a cell?
• Smallest Archae genome from Nanoarchaeum
equtans, 0.491 Mbp: obligate symbiont lacks genes
required for synthesis of lipids, amino acids,
nucleotides, and vitamins, and hence must grow in
close association with another organism which
provides these nutrients.
• Smallest Bacterial genome from Mycoplasma
genitalium, 0.58 Mbp: obligate intracellular pathogen
lacks genes required for amino acid biosynthesis and
the peptidoglycan cell wall, genes encoding TCA
cycle enzymes, and many other biosynthetic genes.
• The smallest free-living organisms have a
genome size over 1 Mbp.
http://www-micro.msb.le.ac.uk/109/Genomes.html
Small molecules of the “average cell”
• Varieties: (1) precursors (building blocks) of
macromolecules; (2) metabolic intermediates; (3)
enzyme cofactors; (4) polyamines bound to DNA
• Approximate number of kinds of small molecules:
(1) 120 amino acids, their precursors and derivatives;
(2) 100 nucleotides, their precursors and derivatives;
(3) 50 fatty acids and their precursors; (4) 250 sugars,
carbohydrates, and their precursors; (5) 300 quinones,
polyisoprenoids, prophyrins, vitamins, other
coenzymes and prosthetic groups, and their
precursors
Study questions 1
• A newly isolated bacterial species is found to have a
single chromosome with a molecular weight of 1 x 109.
Estimate the maximum number of proteinencoding genes it is likely to have if a twodimensional gel reveals that the average molecular
weight of its proteins (polypeptides) is 35,000.
Assume that 10% of the genome does not code for
protein and that 1% codes for stable RNA species.
• Ans: 1,510 (using 110 and 618 as the molecular
weights of an average amino acid residue and an
average nucleotide base-pair residue, respectively)
Study questions 2
• For the reference culture of E. coli described in Table
1, calculate the number of ribosomes in a newborn
cell.
• Ans: 14,060 ribosomes per newborn cell
A giant step toward the creation of artificial
life: synthetic genome
• A team led by Dr. Hamilton Smith, director of the Venter
Institute's Synthetic Biology Group, has manufactured from
laboratory chemicals a ring of DNA containing all the
genes of Mycoplasma genitalium - the tiniest bacteria ever
found. That means the team is tantalizingly close to
creating an artificial form of life that could replicate itself
using these machine-made genes.
• The feat is described in an online edition of the journal
Science released Thursday (January 24, 2008 ) by
researchers at the J. Craig Venter Institute in Rockville, Md.
• The secret to the success of the project was finding ways
to assemble the 100 pieces of genome into subgroups,
then joining the subgroups into successive larger pieces,
until the entire genome could be spliced together from four
lengthy chains.
A giant step toward the creation of artificial
life: transplant of synthetic genome
• In August (2007), the Venter Institute team reported that
they had performed a successful transplant of a natural
genome by removing the chromosome from one
Mycoplasma species and implanting it into another,
which began replicating copies of the first species.
• The plan is to slip the synthetic chromosome inside the
microscopic skin of one of the Mycoplasma bacterium,
replacing its natural genome with the machine-made one
and sparking the creature into a life form that can
reproduce itself.
• There are still several technical hurdles to pass before a
similar procedure could work with the synthetic
chromosome.
A giant step toward the creation of artificial
life: impact
• The work is not merely a demonstration of laboratory
finesse, Venter insisted, but a step toward development
of technologies that could grow fuel in bacterial vats and
speed cures for diseases. "It puts a lot of power in the
hands of humans," he said.
• And there is the matter of bragging rights of mythological
proportions. Mere mortals have yet to lay claim to
creating life.
A giant step toward the creation of artificial
life: next step
• Once the laboratory produces living, replicating bacteria
using this artificial chromosome, Venter scientists plan to
strip away genes systematically, to find how few are truly
necessary to sustain life. It is largely an academic
exercise, but in the process the scientists hope to refine
the tools for building living organisms from this
fundamental base, and custom-design them to perform
certain tasks - such as manufacturing fuel.
A giant step toward the creation of artificial
life: biosafty and other concern
• Jim Thomas, a Montreal researcher for ETC, a Canadian
environmental and social justice advocacy group, said the
"synthetic biology" work pursued by Venter's group is
potentially dangerous and ought to be subject to
government oversight.
• "There are real concerns about biosafety for synthetic
organisms, and this takes us a step closer to them," he
said. "Because of the push toward rapid commercialization,
an environmental release of a synthetic organism is
inevitable. This is an ecological disaster waiting to
happen."
• The push to develop synthetic fuels using these bugs, he
suggested, will place more stress on farmland to produce
energy crops. "We are already seeing fuel versus food
conflicts because of the drive to produce ethanol," he said.