Acholeplasma laidlawii B-PG9 adenine-specific purine nucleoside phosphorylase that accepts ribose-1-phosphate, deoxyribose-1-phosphate, and xylose-1-phosphate

Abstract

An adenylate-specific purine nucleoside phosphorylase (purine nucleoside:orthophosphate ribosyltransferase, EC12.4.2.1) (PNP) was isolated from a cytoplasmic fraction of Acholeplasma laidlawii B-PG9 and partially purified (820-fold). This partially purified PNP could only ribosylate adenine and deribosylate adenosine and deoxyadenosine. The A. laidlawii partially purified PNP could not use hypoxanthine, guanine, uracil, guanosine, deoxyguanosine, or inosine as substrates, but could use ribose-l-phosphate, deoxyribose-1-phosphate, or xylose-l-phosphate as the pentose donor. Mg2e and a pH of 7.6 were required for maximum activity for each of the pentoses. The partially purified enzyme in sucrose density gradient experiments had an approximate molecular weight of 108,000 and a sedimentation coefficient of 6.9, and in gel filtration experiments it had an approximate molecular weight of 102,000 and a Stoke's radius of 4.1 nm. Nondenaturing polyacrylamide tube gels of the enzyme preparation produced one major and one minor band. The major band (R.P 0.57) corresponded to all enzyme activity. The Kms for the partially pwrified PNP with ribose-l-phosphate, deoxyribose-l-phosphate, and xylose-l-phosphate were 0.80, 0.82, and 0.81 mM, respectively. The corresponding V11,,s were 12.5, 14.3, and 12.0 FIM min-', respectively. The Hill or interaction coefficients (n) for all three pentose phosphates were close to unity. The characterization data suggest the possibility of one active site on the enzyme which is equally reactive toward each of the three pentoses. This is the first report of an apparently adenine-specific PNP activity.Purine nucleoside phosphorylase (purine nucleoside:orthophosphate ribosyltransferase, EC 2.4.2

TheMycoplasmas – a congruent path toward minimal life functions

“…In this case, a solution relying on membrane-bound broadaffinity enzymes with 5 -nucleotidase activity that converts nontransportable nucleotides into transportable nucleosides (Pollack, 2002b). Purine nucleoside phosphorylase (PNP) has been shown to have equal activity for nucleobases and nucleosides (McElwain et al ., 1988). The "patchwork" model of metabolic evolution suggests that the evolution of metabolic pathway has been driven by the expansion or alteration of enzyme substrate specificity.…”