Biosynthesis of Proline.

Proline was among the last biosynthetic precursors to have its biosynthetic pathway unraveled. This review recapitulates the findings on the biosynthesis and transport of proline. Glutamyl kinase (GK) catalyzes the ATP-dependent phosphorylation of L-glutamic acid. Purification of γ-GK from Escherichia coli was facilitated by the expression of the proB and proA genes from a high-copy-number plasmid and the development of a specific coupled assay based on the NADPH-dependent reduction of GP by γ-glutamyl phosphate reductase (GPR). GPR catalyzes the NADPH-dependent reduction of GP to GSA. Site directed mutagenesis was used to identify residues that constitute the active site of E. coli GK. This analysis indicated that there is an overlap between the binding sites for glutamate and the allosteric inhibitor proline, suggesting that proline competes with the binding of glutamate. The review also summarizes the genes involved in the metabolism of proline in E. coli and Salmonella. Among the completed genomic sequences of Enterobacteriaceae, genes specifying all three proline biosynthetic enzymes can be discerned in E. coli, Shigella, Salmonella enterica, Serratia marcescens, Erwinia carotovora, Yersinia, Photorhabdus luminescens, and Sodalis glossinidius strain morsitans. The intracellular proline concentration increases with increasing external osmolality in proline-overproducing mutants. This apparent osmotic regulation of proline accumulation in the overproducing strains may be the result of increased retention or recapture of proline, achieved by osmotic stimulation of the ProP or ProU proline transport systems. A number of proline analogs can be incorporated into proteins in vivo or in vitro.