Purification and characterization of a molybdenum-pterin-binding protein (Mop) in Clostridium pasteurianum W5.

A large-scale fractionation scheme purified the major molybdenum(Mo)-binding protein (Mop) from crude extracts of Clostridium pasteurianum, with a 10 and 0.2% yield of Mo and protein, respectively. The apparent molecular weight of the purified molybdoprotein is 5,700, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The protein contains 0.7 mol of Mo per mol of protein with a molecular weight of 5,700. Mop, as isolated, has a peak absorbency at 293 nm. Denaturation and oxidation of the molybdoprotein released multiple pterin like fluorescent compounds. Mop appears to contain a pterin derivative and Mo, but phosphate analysis indicated that the pterin at the very least is not phosphorylated; phosphorylation is required for functional molybdenum cofactor. All treatments used to release the putative Mo-pterin species from Mop failed to yield a molybdopterin that had detectable molybdenum cofactor activity.