Abstract
The role of aspartate-84, an invariant residue in the active site cleft of Escherichia coli por-
phobilinogen deaminase, has been investigated by site-directed mutagenesis. Substitution of aspartate-84 by glutamate results in an enzyme that retains less than 1% of its activity and which can form highly stable
enzyme-intermediate complexes. Substitution of aspartate-84 by either alanine or asparagine, however, results in proteins unable to catalyze the formation of preuroporphyrinogen but which, nevertheless, appear abletoassemblethedipyrromethanecofactor. Themechanismsofthetetramerizationreactionandcofactor assembly are discussed.