INVOLVEMENT OF THE C-TERMINUS IN INTRAMOLECULAR NITROGEN CHANNELING IN GLUCOSAMINE 6-PHOSPHATE SYNTHASE - EVIDENCE FROM A 1.6 ANGSTROM CRYSTAL-STRUCTURE OF THE ISOMERASE DOMAIN

Background: Glucosamine 6-phosphate synthase (GlmS) catalyses the firs t step in hexosamine metabolism, converting fructose-6P (6 phosphate) into glucosamine-6P using glutamine as a nitrogen source. GlmS is a bi enzyme complex consisting of two domains that catalyse glutamine hydro lysis and sugar-phosphate isomerisation, respectively. Knowledge of th e three-dimensional structure of GlmS is essential for understanding t he general principles of catalysis by ketol isomerases and the mechani sm of nitrogen transfer in glutamine amidotransferases, Results: The c rystal structure of the isomerase domain of the Escherichia coli GlmS with the reaction product, glucosamine-6P, has been determined at 1.57 Angstrom resolution. It is comprised of two topologically identical s ubdomains, each of which is dominated by a nucleotide-binding motif of a flavodoxin type. The catalytic site is assembled by dimerisation of the protein. Conclusions: The isomerase active site of GlmS seems to be the result of evolution through gene duplication and subsequent dim erisation. Isomerisation of fructose-6P is likely to involve the forma tion of a Schiff base with Lys603 of the enzyme, the ring-opening step catalysed by His504, and the proton transfer from C1 to C2 of the sub strate effected by Glu488. The highly conserved C-terminal fragment of the chain may play a key role in substrate binding, catalysis and com munication with the glutaminase domain. The corresponding sequence pat tern DXPXXLAK[SC]VT tin single-letter aminoacid code, where X is any a mino acid and letters in brackets indicate that either serine or cyste ine may take this position) may be considered as a fingerprint of GlmS .