Crystal structures of Salmonella typhimurium propionate kinase and its complex with Ap₄A: Evidence for a novel Ap₄A synthetic activity

Abstract

Propionate kinase catalyses the last step in the anaerobic breakdown of L-threonine to propionate in which propionyl phosphate and ADP are converted to propionate and ATP. Here we report the structures of propionate kinase (TdcD) in the native form as well as in complex with diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) by X-ray crystallography. Structure of TdcD obtained after cocrystallization with ATP showed Ap4A bound to the active site pocket suggesting the presence of Ap4A synthetic activity in TdcD. Binding of Ap4A to the enzyme was confirmed by the structure determination of a TdcD-Ap4A complex obtained after cocrystallization of TdcD with commercially available Ap4A. Mass spectroscopic studies provided further evidence for the formation of Ap4A by propionate kinase in the presence of ATP. In the TdcD-Ap4A complex structure, Ap4A is present in an extended conformation with one adenosine moiety present in the nucleotide binding site and other in the proposed propionate binding site. These observations tend to support direct in-line transfer of phosphoryl group during the kinase reaction.

Structure and function of enzymes involved in the anaerobic degradation of L-threonine to propionate

“…Our studies on propionate kinase from S. typhimurium provided evidence for a novel diadenosine 5′,5′′′-P 1 ,P 4tetraphosphate (Ap 4 A) synthetic activity (Simanshu et al 2007). Crystals of TdcD obtained in the presence of ATP clearly showed Ap 4 A bound in the active site pocket of the enzyme.…”

Structural and mechanistic investigations on Salmonella typhimurium acetate kinase (AckA): identification of a putative ligand binding pocket at the dimeric interface

“…As AckA is involved at the junction of carbohydrate and fatty acid metabolism, it might be a suitable target for development of inhibitors [34]. We have earlier structurally and biochemically characterized Salmonella typhimurium propionate kinase ( St TdcD) and demonstrated that the enzyme is specific to propionate although could utilize acetate at a 10 times lower rate [13,35] . In the present study, we report structural and mechanistic investigations of St AckA.…”

What is the conformation of physiologically-active dinucleoside polyphosphates in solution? Conformational analysis of free dinucleoside polyphosphates by NMR and molecular dynamics simulations

“…18 Crystal structures of several kinases or hydrolases-bound dinucleotides in enzymes were determined. [19][20][21][22] In protein-bound dinucleotides, the polyphosphate chain adopts various conformations: extended, 19-22 S-shaped 21 or folded. 22 In some cases the proteinbound dinucleotides coordinate with a Mg 2+ ion.…”