Purification and crystallization of precursors and autoprocessed enzymes of Flavobacterium glycosylasparaginase: an N-terminal nucleophile hydrolase

Glycosylasparaginase (GA) represents a novel group of proteins that are act ivated by self-catalyzed peptide-bond cleavage from a single-chain precurso r to yield the two subunits required for hydrolase activity. The wild-type GA precursor autoproteolyzes spontaneously into alpha and beta subunits. St rategies are reported here for purification to homogeneity of GA from Flavo bacterium meningosepticum in both single-chain precursor and mature (autopr ocessed) forms. The recombinant proteins crystallize in different space gro ups: P1 and P2(1) for the precursor and mature enzymes, respectively. The p recursor crystals diffract to 1.9 Angstrom resolution with laboratory X-ray radiation.