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#ATPase

5 public questions tagged with this topic.

Membrane Transportation

Which ATPase is crucial for acidifying vacuoles in plant cells?

Plant central vacuole occupies major cellular volume functioning in turgor generation, storage of ions, sugars, pigments, anthocyanins and toxic metabolites, pH regulation and degradation analogous to animal lysosomes. Luminal pH typically 5 to 5.5 maintained acidic relative to cytosol 7.5 to support proton coupled antiport of calcium via CAX, sodium via NHX, sugars and sequestration of xenobiotics. Acidification achieved by two proton pumps coexisting in tonoplast: V-type proton ATPase and proton pyrophosphatase using pyrophosphate. Dominant contributor is V-ATPase, multisubunit rotary machine similar to lysosomal pump, comprising V1 domain A3B3 hexamer hydrolyzing ATP and Vo domain containing c-ring proteolipids and subunit a forming proton path, consuming cytosolic ATP to move protons inward without phosphoenzyme intermediate, sensitive to bafilomycin and concanamycin. Pyrophosphatase provides backup using PPi from metabolism. F-type ATP synthase located exclusively in mitochondria chloroplasts synthesizes ATP driven by proton motive force, does not reside in vacuole. Na+/K+ ATPases absent in higher plants; calcium ATPases distinct. Thus plant vacuolar acidification crucially depends on V-ATPase creating proton motive force.

Ref: Sze et al., Trends Plant Sci, Plant Vacuolar V-ATPase Acidification of Vacuoles and Tonoplast.

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Cytoskeleton

The AAA+ domain in dynein is responsible for:

Structural analysis of dynein heavy chain shows tail responsible for dimerization and cargo adaptor binding, followed by six AAA+ domains arranged as heterohexameric ring with central pore. AAA1 bears conserved Walker A P-loop GXXXXGKT and Walker B hhhhDE motif essential for ATP binding and hydrolysis, AAA2-AAA4 modulate allosteric communication, AAA5-AAA6 stabilize ring and interact with stalk and strut. Upon ATP hydrolysis at AAA1, conformational changes propagate around ring altering interface between AAA5 and stalk buttress, shifting coiled-coil registry and linker docking. This generates measurable force of several piconewtons per stroke. Mutagenesis of lysine in Walker A or glutamate in Walker B abolishes motility, dominant negative in cells. Cargo attachment mediated by N-terminal tail associating with intermediate chain, light intermediate chain, light chains LC8, Tctex, Roadblock that bind adaptors like BICD, HOOK, Spindly. No interaction with actin filaments, no direct tubulin polymerization regulation, distinguishing AAA ring as force generating engine analogous to other ring-translocases adapted for cytoskeletal motor function.

Ref: Carter et al. Nature 2011; AAA+ AAA1 Walker A/B hydrolyzes ATP generating 4-5 pN force.

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Cytoskeleton

Myosin head movement along actin filaments is driven by:

Myosin motor cycle couples ATP hydrolysis to filament sliding via conformational changes in conserved motor domain. In absence of ATP myosin head strongly bound to actin in rigor state, stereospecific binding between actin helix and myosin cardiomyopathy loop. Binding of ATP to P loop pocket between upper and lower 50 kilodalton subdomains induces opening of actin binding cleft reducing affinity thousand fold leading to detachment. While detached ATP hydrolyzed to ADP Pi mediated by Switch I Switch II closing around gamma phosphate, triggering recovery stroke where lever arm moves about 90 degrees to pre power stroke conformation storing elastic energy. Myosin ADP Pi rebinds actin weakly, phosphate release gates transition to strongly bound state closing cleft, producing power stroke large rotation of converter domain swinging lever arm 5 to 10 nanometer dragging actin. ADP release returns to rigor awaiting new ATP. Thus detachment powered by ATP binding, priming by hydrolysis, force generation by Pi release, not calcium influx, GTP hydrolysis, tail phosphorylation primary driver of movement.

Ref: Alberts et al., Molecular Biology of the Cell, 7th ed., Chapter 16: Myosin ATP Hydrolysis and Movement.

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Protein Sorting and Trafficking

Which ATPase is required for post-translational protein translocation into the ER?

While most secretory and membrane proteins in mammals rely on SRP-dependent co-translational targeting, small proteins less than about one hundred residues and many soluble yeast proteins synthesized completely before targeting rely on post-translational translocation requiring distinct chaperone system. Cytosolic Hsp70 homologs Ssa1 maintain substrate unfolded, then Sec62/Sec63 heterodimer together with Sec71/Sec72 accessory proteins in yeast forms heptameric Sec complex around Sec61 channel to recruit substrate. Lumenal Hsp70 BiP, called Kar2 in Saccharomyces, binds DnaJ domain of Sec63 which stimulates its ATPase activity. ATP-bound BiP recruited to incoming chain hydrolyzes ATP to ADP, converting to closed high-affinity conformation clamping onto hydrophobic patches, preventing back-sliding through channel. Nucleotide exchange factors Sil1 and Grp170 promote ADP release for BiP recycling, allowing repeated capture events that rectify Brownian motion into directional pulling force. Each ATP hydrolysis cycle advances chain inward independent of ribosome. SRP, Sec61 alone without BiP and Ran-GTP dependent nuclear import do not provide ATP-driven pulling for this mode, so BiP is essential ATPase converting chemical energy into vectorial movement for post-translational ER entry and secretory pathway maintenance.

Ref: Matlack et al., Cell 97: 1999, BiP and Sec63 in Post-translational Translocation.

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