Role of novel dye-linked dehydrogenases in the metabolism of polyethylene glycol by pure cultures ofSphingomonassp. N6

Abstract

Abstract Polyethylene glycol and diglycolic acid dehydrogenase acitivities linked with3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2 H-tetrazolium bromide and phenazine methosulfate were found in the participate fraction of the sonic extract of a newly isolated polyethylene glycol 20 000-utilizing bacterium ( Sphingomonas sp. N6). The amount of glyoxylic acid formed from diglycolic acid increased proportionally with the increase in reaction time and enzyme concentration to show that diglycolic acid can be a model compound for an ether-cleaving enzyme. Both enzymes were formed inducibly when the organism was grown on polyethylene glycol 10000. Both enzymes transferred many more electrons to3-(4,5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2 H-tetrazolium bromide plus phenazine methosulfate than to 2,6-dichlorophenolindophenol plus phenazine methosulfate. Also, ferricyanide, nitroblue tetrazolium and horse heart cytochrome c served as electron acceptors, among which3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2 H-tetrazolium bromide plus phenazine methosulfate was most active for polyethylene glycol dehydrogenase and ferricyanide was most active for diglycolic acid dehydrogenase irrespective of the presence of phenazine methosulfate. NAD and NADP did not act as electron acceptors. Polyethylene glycol dehydrogenase was completely inhibited by 1,4-benzoquinone and partially inhibited by quinine and glyoxylic acid. Diglycolic acid dehydrogenase was strongly inhibited by 1,4-benzoquinone and partially inhibited by α-benzoin oxime, quinacrine and glyoxylic acid. The enzymes appear to be different from each other and also from polyethylene glycol and diglycolic acid dehydrogenases of polyethylene glycol 20 000-utilizing symbiotic mixed culture E-1in which S. terrae was responsible for polyethylene glycol degradation in the coupling with electron acceptors and in the effect of inhibitors.