Phosphoribulokinase: Current perspectives on the structure/function basis for regulation and catalysis

Phosphoribulokinase (PRK), an enzyme unique to the reductive pentose phosphate pathway of CO2 assimilation, exhibits distinctive contrasting properties when the proteins from eukaryotic and prokaryotic sources are compared. The eukaryotic PRKs are typically dimers of -39 kDa subunits while the prokaryotic PRKs are octamers of -32 kDa subunits. The enzymes from these two classes are regulated by different mechanisms. Thioredoxin f mediated thiol-disulfide exchange interconverts eukaryotic PRKs between reduced (active) and oxidized (inactive) forms. Allosteric effectors, including activator NADH and inhibitors AMP and phosphoenolpyruvate, regulate activity of prokaryotic PRK. The effector binding site has been identified in the high resolution structure recently elucidated for prokaryotic PRK and the apparatus for transmission of the allosteric stimulus has been identified. Additional contrasts between PRKs include marked differences in primary structure between eukaryotic and prokaryotic PRKs. Alignment of all available deduced PRK sequences indicates that less that 10% of the amino acid residues are invariant. In contrast to these differences, the mechanism for ribulose 1,5-bisphosphate synthesis from ATP and ribulose 5-phosphate (Ru5P) appears to be the same for all PRKs. Consensus sequences associated with M++-ATP binding, identified in all PRK proteins, are closely juxtaposed to the residue proposed to function as general base catalyst. Sequence homology and mutagenesis approaches have suggested several residues that may potentially function in Ru5P binding. Not all of these proposed Ru5P binding residues are closely juxtaposed in the structure of unliganded PRK. Mechanistic approaches have been employed to investigate the amino acids which influence K-m Ru5P and identify those amino acids most directly involved in Ru5P binding. PRK is one member of a family of phospho or sulfo transferase proteins which exhibit a nucleotide monophosphate kinase fold. Structure/function correlations elucidated for PRK, suggest analogous assignments for other members of this family of proteins.