ACTIVE-SITE STRUCTURE OF METHYLAMINE DEHYDROGENASE - HYDRAZINES IDENTIFY C6 AS THE REACTIVE SITE OF THE TRYPTOPHAN-DERIVED QUINONE COFACTOR

To identify the reactive part of the ortboquinone function of the tryptophan-derived cofactor found in methylamine dehydrogenase (MADH), we have determined the crystal structures of MADH from Thiobacillus versutus inhibited by methylhydrazine and (2,2,2-trifluoroethyl)hydrazine. Extra electron density attached to C6 of the tryptophyl tryptophanquinone cofactor shows that this atom and not C7 is the reactive part of the ortho-quinone moiety. The density retained after hydrazine inhibition is much less extensive than expected, however, suggesting that partial breakdown of the inhibitors after reaction with the cofactor may take place. A detailed description is presented of the cofactor environment in an improved model of MADH which now includes information from the recently determined gene sequence of the cofactor-containing subunit [Ubbink, M., van Kleef, M. A. G., Kleinjan, D., Hoitink, C. W. G., Huitema, F., Beintema, J.J., Duine, J.A., & Canters, G.W.(1991) Eur. J. Biochem. 202, 1003-1012]. We hypothesize that Asp76 is responsible for proton abstraction from the alpha-carbon of the substrate during catalysis.