Microbial oxidation of methane and methanol: purification and properties of a heme-containing aldehyde dehydrogenase from Methylomonas methylovora.
Procedures for the purification of an aldehyde dehydrogenase from extracts of the obligate methylotroph, Methylomonas methylovora are described. The purified enzyme is homogeneous as judged from polyacrylamide gel electrophoresis. In the presence of an artificial electron acceptor (phenazine methosulfate), the purified enzyme catalyzes the oxidation of straight chain aldehydes (C1--C10 tested), aromatic aldehydes (benzaldehyde, salicylaldehyde), glyoxylate, and glyceraldehyde. Biological electron acceptors such as NAD+, NADP+, FAD, FMN, pyridoxal phosphate, and cytochrome c cannot act as electron carriers. The activity of the enzyme is inhibited by sulfhydryl agents [p-chloromercuribenzoate, N-ethylmaleimide and 5,5-dithiobis (2-nitrobenzoic acid)], cuprous chloride, and ferrour nitrate. The molecular weight of the enzyme as estimated by gel filtration is approximately 45000 and the subunit size determined by sodium dodecyl sulfate-gel electrophoresis is approximately 23000.  The purified enzyme is light brown and has an absorption peak at 410 nm. Reduction of enzyme with sodium dithionite or aldehyde substrate resulted in the appearance of peaks at 523 nm and 552nm. These results suggest that the enzyme is a hemoprotein. There was no evidence that flavins were present as prosthetic group. The amino acid composition of the enzyme is also presented.
D000445:Aldehyde Oxidoreductases / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D002474:Cell-Free System; D006418:Heme / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D008697:Methane / Q000378:metabolism*; D000432:Methanol / Q000378:metabolism*; D008744:Methylamines / Q000378:metabolism; D008772:Methylococcaceae / Q000201:enzymology* / Q000378:metabolism; D008970:Molecular Weight; D013379:Substrate Specificity; D013696:Temperature
D000596:Amino Acids; D008744:Methylamines; D006418:Heme; D000445:Aldehyde Oxidoreductases; D008697:Methane; D000432:Methanol