Prokaryote Biology Module 1

Who overturned the concept of the prokaryote, and with what tree?

Carl Woese; the three-domain tree (Woese and Fox, 1977).

What molecule did Woese use for taxonomic classification?

Ribosomal RNA (rRNA).

Why can rRNA be used to compare all organisms?

It is fundamental to all living organisms.

Which rRNA gene is sequenced in prokaryotes vs. eukaryotes?

16S in prokaryotes; 18S in eukaryotes.

What method did Woese use before rRNA gene sequencing existed?

rRNA "fingerprinting."

What isotope labeled the rRNA in fingerprinting?

Radioactive P32, supplied in the growth media.

After which base was the labeled rRNA cleaved?

Guanine.

How were the rRNA fragments separated and compared?

Homochromatography by charge and base composition, comparing migration patterns of shared fragments.

What organisms did Ralph Wolfe study that led toward the discovery of Archaea?

Methanogens, whose biochemistry is unusual.

What did Wolfram Zillig find about archaeal RNA polymerase?

The basal machinery of Archaea and Eukarya is related.

What two structural distinctions did Otto Kandler document?

Phospholipid and cell wall differences; all archaea lack peptidoglycan.

Why did Karl Stetter's hyperthermophile isolations matter?

They provided a greater diversity of microorganisms for classification.

Which domain branches first in the three-domain tree?

Bacteria.

In the three-domain tree, which domain is sister to Eukarya?

Archaea.

What kind of organisms occupy the deepest branches of the tree?

Hyperthermophiles, roughly 80 C anaerobes, in both Bacteria and Archaea.

Was LUCA aerobic or anaerobic, and how heat-loving?

Definitely an anaerobe; a moderate thermophile or mesophile.

In what setting is LUCA proposed to have lived, and what supplied its gradient?

A white smoker; serpentinization, which also forms formate.

What metabolism is proposed for LACA?

Methane and short-chain alkane metabolism.

What does the two-domain tree claim about eukaryotes?

They arise from within Archaea rather than as a separate domain.

In the mitochondria-late model, what engulfs what?

An advanced eukaryote already capable of phagocytosis engulfs an alpha-proteobacterium.

Where were the Asgard archaea discovered?

Loki's Castle.

What does ESP stand for, and give two examples.

Eukaryotic signature protein; ubiquitin-like proteins and actin.

What morphological features does Lokiarchaeota strain MK-D1 have?

Tentacle-like protrusions, a complex cell wall, and an actin filament network.

What does MK-D1 produce, and which partners consume it?

Formate and H2, used by sulfate-reducing bacteria and methanogens.

What energizes rotation of the bacterial flagellum?

Proton motive force, or sodium motive force in some bacteria.

What is the diameter of the bacterial flagellar filament, and what anchors it?

Roughly 10-20 nm; basal body rings.

The bacterial flagellum is evolutionarily related to what?

The type III secretion system.

What is a lophotrichous arrangement?

A patch of flagella at one end of the cell.

What is a peritrichous arrangement? Give an example.

Flagella over the whole surface; E. coli.

What is a polar arrangement? Give an example.

A single flagellum at one pole; Vibrio.

What happens when flagella rotate counterclockwise?

They bundle, and the cell swims smoothly.

What happens when flagella rotate clockwise?

The bundle disperses and the cell tumbles.

What determines which rotation direction dominates?

Attractant and repellent cues.

Which domain has the fastest swimmers, and using what?

Archaea, using the archaellum.

What is the archaellum's filament diameter, and what flagellar part does it lack?

About 12 nm; it has no basal body rings.

What energizes the archaellum?

ATP hydrolysis, not an ion gradient.

The archaellum is evolutionarily related to what?

Type IV pili.

What dynamic behavior defines type IV pili, and what does it let the cell do?

Polymerization and depolymerization; they act as the cell's sense of touch and initiate biofilms.

What enzymes drive type IV pilus extension and retraction?

Assembly and retraction ATPases.

How do fimbriae compare to type IV pili, and what do their tips bind?

Much shorter and more numerous; they bind oligosaccharides on glycoproteins and glycolipids.

What do conjugation (F) pili do, and does DNA pass through them?

They join donor to recipient; DNA passes through the cell-to-cell contact, not the pilus.

How does a slime layer differ from a glycocalyx, and what counts as glycocalyx?

Slime is loosely attached, glycocalyx tightly attached; capsules and S-layers.

Name four functions of the glycocalyx.

Adhesion, biofilm formation, protection from phagocytosis, and desiccation resistance.

What is an S-layer made of?

A rigidly attached array of protein or glycoprotein.

How can an S-layer be attached in archaea?

Integral to the membrane, lipid-anchored, or bound to pseudopeptidoglycan.

Where does the S-layer sit in bacteria?

On the surface of the cell wall.

Under what conditions are nanowires produced, and what carries the electrons?

Low agitation with a limiting electron acceptor such as oxygen; chains of heme cytochromes.

What do cellulosomes do?

Hydrolyze cellulose at the cell surface, releasing sugars.

What are archaeal cannulae?

Tubes connecting the periplasmic space of neighboring archaeal cells.

What does the archaeal hamus look like, and what is it for?

Barbed wire with a grappling hook end; attachment and biofilm formation.

What linkage joins tails to glycerol in archaea vs. bacteria and eukaryotes?

Ether in archaea; ester in bacteria and eukaryotes.

Compare the lipid tails of archaea and bacteria.

Archaea use branched isoprenoid chains; bacteria use unbranched fatty acids, saturated or unsaturated.

Which glycerol stereoisomer does each group use?

L-glycerol in archaea; D-glycerol in bacteria and eukaryotes.

Why can archaea build a membrane monolayer?

Tetraether lipids span the entire membrane.

What backbone do sphingolipids use, and where are they found?

Sphingosine; in some bacteria and in mammals.

About what fraction of the membrane is protein, and how do those proteins attach?

Roughly 60 percent; integral, peripheral, or covalently lipid-anchored.

Name four functions of cytoplasmic membrane proteins.

Generating gradients, nutrient transport, environmental signaling, and anchoring structures such as the flagellum.

What do sterols do, and which prokaryotes have them?

Strengthen the membrane; Thermoplasma and Mycoplasma.

What sugars make up peptidoglycan, how are they linked, and what is odd about the cross bridges?

NAG and NAM, beta-1,4 linked; the cross bridges use both L- and D-amino acids.

What enzyme cleaves peptidoglycan?

Lysozyme.

Where is peptidoglycan located in Gram-positive vs. Gram-negative cells?

A thick layer outside the membrane; a thin layer in the periplasm.

How does cross-linking differ between Gram-negative and high-GC Gram-positive bacteria?

Gram-negatives link directly; high-GC Gram-positives use an amino acid bridge, making the wall more rigid.

What are teichoic acids made of, and why are they strongly negative?

Ribitol phosphate or glycerol phosphate polymers; their repeating phosphates carry the charge.

Name two functions of teichoic acids.

Stabilizing the cell wall and providing attachment sites for enzymes and phage receptors.

Which Gram-positive lineage lacks peptidoglycan entirely?

Mycoplasma (mollicutes), which therefore do not gram stain.

What defines an acid-fast bacterium, and name one.

Resistance to decolorization by acid alcohol; the tuberculosis agent.

What lies above the peptidoglycan in an acid-fast envelope, and what is interdigitated with it?

The mycomembrane with mycolic acids above; arabinogalactan interdigitated with the peptidoglycan.

What occupies the inner and outer leaflets of the Gram-negative outer membrane?

Phospholipid inside; lipopolysaccharide (LPS) outside.

Name three functions of LPS.

Barrier against hydrophobic compounds, stabilizing surface structures by binding Mg2+, and triggering the immune system.

What protein fold dominates the outer membrane, and what is a porin?

Beta barrels; a porin is a triple beta barrel channel for hydrophilic solutes.

Name the non-specific porins and two substrates with specific porins.

OmpC, OmpF and PhoE; maltose (LamB) and sucrose.

Why is there no active transport across the outer membrane?

It has neither ATP nor an electrochemical gradient.

What does OmpA do?

Maintains outer membrane stability and acts as a receptor for phage, peptides, and the F-pilus.

Name the roles of murein lipoprotein, OmpX, and OmpLA.

Murein lipoprotein anchors the outer membrane to peptidoglycan; OmpX is a virulence protein; OmpLA is a destabilizing phospholipase.

Name three periplasmic processes, and two things the periplasm lacks.

Redox reactions, disulfide bond formation, and hydrolysis; it lacks ATP and an electrochemical gradient.

How are large molecules imported across the outer membrane, and how is that powered?

TonB-dependent transport, powered by the inner membrane gradient via the ExbB/ExbD complex.

What holds for all archaeal cell walls, and is the gram stain informative?

All lack peptidoglycan; the gram stain does not correlate with archaeal phylogeny.

How does pseudopeptidoglycan differ from peptidoglycan?

NAT replaces NAM, linkages are beta-1,3, and only L-amino acids are used.

Which archaeon has an outermost membrane, and what associates with it?

Ignicoccus, which has a huge periplasm; Nanoarchaeum associates with it.

What is the packaged bacterial chromatin structure called, and what maintains it?

The nucleoid, maintained by nucleoid-associated proteins.

What are the two classes of nucleoid-associated protein, and what does each do?

Bridgers hold separate DNA segments together; benders fold the chromatin.

How do archaea package their chromatin?

With histone-like proteins, though these are not universally conserved.

What does LLPS stand for, and what does it produce?

Liquid-liquid phase separation; transient compartments that are not membrane bound.

What interactions drive LLPS?

Cation-anion, dipole-dipole (pi-pi), and cation-pi interactions.

What are BR bodies made of, what do they do, and what enzyme drives their formation?

RNA and protein; they organize mRNA decay; RNase E.

What is the purpose of intracytoplasmic membranes, and what shapes do they take?

To increase surface area for biochemistry; tubes, vesicles, and lamellae.

What do the ICMs of methanotrophs and nitrogen fixers accomplish?

Methanotrophs house methane oxidation enzymes; nitrogen fixers raise respiration, removing oxygen and supplying ATP for nitrogenase.

What is unusual about the Planctomycete cell wall?

It is proteinaceous and lacks peptidoglycan.

What distinguishes the pirellulosome from the paryphoplasm?

The pirellulosome contains ribosomes; the paryphoplasm lacks them.

What reaction occurs in the anammoxosome, and what toxic intermediate does it sequester?

Anaerobic oxidation of ammonia to N2 using nitrite as electron acceptor; hydrazine.

What is a chlorosome made of, and which organisms have one?

A galactolipid monolayer packed with photopigments; green photosynthetic bacteria.

Where are phycobilisomes located, and in which organisms?

On photosystem II of the thylakoid membrane; cyanobacteria and red algae.

What minerals form magnetosomes, and what do they do?

Magnetite and greigite; they orient the cell along geomagnetic field lines.

What encloses a PHB granule, and what is PHB useful as?

A phospholipid monolayer; a biodegradable bioplastic.

How do polyphosphate granules and acidocalcisomes differ?

Polyphosphate granules form spontaneously, unbound by lipid; acidocalcisomes are lipid bound and mineral rich.

What are gas vesicles made of, and what do they do?

Protein only, from a coat encoded by a single gene; they allow flotation and raise the surface area to volume ratio.

What controls gas vesicle formation, and what happens once one collapses?

Light; a collapsed vesicle cannot be resynthesized.

What coats a polyhedral organelle, and what does the carboxysome do?

Protein rather than membrane; the carboxysome concentrates CO2 for RuBisCO.

Where are the active sites of self-compartmentalizing proteases, and why do they need energy?

Buried inside the complex; an ATPase must unfold the substrate and feed it in.

How are the smaller self-compartmentalizing proteases described?

As viroid-like.