ALTERATION OF THE QUATERNARY STRUCTURE OF GLUTAMATE-DEHYDROGENASE FROM CLOSTRIDIUM-SYMBIOSUM BY A SINGLE MUTATION DISTANT FROM THE SUBUNIT INTERFACES

X-ray crystallographic studies have previously shown that glutamate de hydrogenase from Clostridium symbiosum is a homohexamer. Mutation of t he active-site aspartate-165 to histidine causes an alteration in the structural properties of the enzyme. The mutant enzyme, D165H exists p redominantly as a single species of lower molecular mass than the wild -type enzyme as indicated by gel filtration and sedimentation velocity analysis. The latter technique gives an s(20,w) value for D165H of (6 .07+/-0.01)S which compares with (11.08+/-0.01)S for the wild-type, in dicative of alteration of the homohexameric quaternary structure of th e native enzyme to a dimeric form, a result confirmed by sedimentation equilibrium experiments. Further support for this is provided by chem ical modification by Ellman's reagent of cysteine-144 in the mutant, a residue which is buried at the dimer-dimer interface in the wild-type enzyme and is normally inaccessible to modification. The results sugg est a possible structural route for communication between the active s ites and subunit interfaces which may be important for relaying signal s between subunits in allosteric regulation of the enzyme.