EVIDENCE FOR THE PRESENCE OF 2 PYRIDINE NUCLEOTIDE-BINDING SITES ON THE BETA-SUBUNIT OF THE ESCHERICHIA-COLI PYRIDINE-NUCLEOTIDE TRANSHYDROGENASE

The pyridine nucleotide transhydrogenase of Escherichia coli is compos ed of two types of subunits, alpha and beta. Trypsin digestion of the purified enzyme generates fragments of the a subunit. The beta subunit is uncleaved unless NADP(H) is present (Tong, R.C.W., Glavas, N.A. an d Bragg, P.D. (1991) Biochim. Biophys. Acta 1080, 19-28). Purified tra nshydrogenase bound to either NAD- or NADP-agarose was treated with tr ypsin. The alpha subunit was cleaved to 16, 29 and 43 kDa fragments in both cases. The beta subunit remained bound to NAD-agarose but was re leased as two cleavage fragments (25 and 30 kDa) from NADP-agarose. Th e beta subunit of the transhydrogenase bound to NAD-agarose was cleave d by trypsin in the presence of NADP(H) to yield 25 and 30 kDa fragmen ts of the beta subunit. These results suggest that the beta subunit co ntains two pyridine nucleotide-binding sites.