1gmh
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REFINED CRYSTAL STRUCTURE OF "AGED" AND "NON-AGED" ORGANOPHOSPHORYL CONJUGATES OF GAMMA-CHYMOTRYPSIN
OverviewOverview
"Aged" organophosphoryl conjugates of serine hydrolases differ from the, corresponding "non-aged" conjugates in their striking resistance to, nucleophilic reactivation. The refined X-ray structures of "aged" and, "non-aged" organophosphoryl conjugates of gamma-chymotrypsin were compared, in order to understand the molecular basis for this resistance of "aged", conjugates. "Aged" and "non-aged" crystalline, organophosphoryl-gamma-chymotrypsin conjugates were obtained by prolonged, soaking of native gamma-chymotrypsin crystals with appropriate, organophosphates. Thus, a representative "non-aged" conjugate, diethylphosphoryl-gamma-chymotrypsin, was obtained by soaking native, crystals with paraoxon (diethyl-p-nitrophenyl phosphate), and a closely, related "aged" conjugate, monoisopropyl-gamma-chymotrypsin, was obtained, by soaking with diisopropylphosphorofluoridate. In both crystalline, conjugates, the refined structures clearly reveal a high occupancy of the, active site by the appropriate organophosphoryl moiety within covalent, bonding distance of Ser195 O gamma. Whereas in the "non-aged" conjugate, both ethyl groups can be visualized clearly, in the putative "aged", conjugate, as expected, only one isopropyl group is present. There is, virtually no difference between the "aged" and "non-aged" conjugates, either with respect to the conformation of the polypeptide backbone as a, whole or with respect to the positioning of the side-chains within the, active site. In the "aged" conjugate, however, close proximity (2.6 A) of, the negatively charged phosphate oxygen atom of the dealkylated, organophosphoryl group to His57 N epsilon 2 indicates the presence of a, salt bridge between these two moieties. In contrast, in the "non-aged", conjugate the DEP moiety retains its two alkyl groups; thus, lacking a, negative oxygen atom, it does not enter into such a charge-charge, interaction and its nearest oxygen atom is 3.6 A away from His57 N epsilon, 2. It is suggested that steric constraints imposed by the salt bridge in, the "aged" conjugate lie at the basis of its resistance to reactivation.
About this StructureAbout this Structure
1GMH is a Protein complex structure of sequences from Bos taurus with SO4 and ISP as ligands. Active as Chymotrypsin, with EC number 3.4.21.1 Full crystallographic information is available from OCA.
ReferenceReference
Refined crystal structures of "aged" and "non-aged" organophosphoryl conjugates of gamma-chymotrypsin., Harel M, Su CT, Frolow F, Ashani Y, Silman I, Sussman JL, J Mol Biol. 1991 Oct 5;221(3):909-18. PMID:1942036
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