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Publication Abstract from PubMed

The homologous human ABO(H) A and B blood group glycosyltransferases GTA and GTB have two mobile polypeptide loops surrounding their active sites that serve to allow substrate access and product egress and to recognize and sequester substrates for catalysis. Previous studies have established that these enzymes can move from the 'open' state to the 'semi-closed' then 'closed' states in response to addition of substrate. The contribution of electrostatic interactions to these conformational changes has now been demonstrated by the determination at various pH of the structures of GTA, GTB, and the chimeric enzyme ABBA. Near neutral pH, GTA displays the closed state in which both mobile loops order around the active site, whereas ABBA and GTB display the open state. At low pH the apparent protonation of the DXD motif in GTA leads to expulsion of the donor analog to yield the open state, while at high pH both ABBA and GTB form the semi-closed state in which the first mobile loop becomes an ordered alpha-helix. Step-wise deprotonation of GTB in increments of 0.5 between pH 6.5 and 10.0 shows that helix ordering is gradual, which indicates that the formation of the semi-closed state is dependent on electrostatic forces consistent with the binding of substrate. Spectropolarimetric studies of the corresponding stand-alone peptide in solution reveal no tendency towards helix formation from pH 7.0 to 10.0, which shows that pH-dependent stability is a product of the larger protein environment and underlines the importance of substrate in active site ordering.

pH-induced conformational changes in human ABO(H) blood group glycosyltransferases confirm the importance of electrostatic interactions in formation of the semi-closed state.,Johal AR, Blackler RJ, Alfaro JA, Schuman B, Borisova S, Evans SV Glycobiology. 2013 Nov 20. PMID:24265507^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.