Amylose chain behavior in an interacting context. III. Complete occupancy of the AMY2 barley alpha-amylase cleft and comparison with biochemical data

In the first two papers of this series the tools necessary to evaluate subs trate ring deformations were developed, and then the modeling of short amyl ose fragments (maltotriose and maltopentaose) inside the catalytic site of barley alpha-amylase was performed In this third paper, this docking has be en extended to the whole catalytic cleft. A systematic approach to extend t he substrate was used on the reducing side from the previous enzyme/pentasa ccharide complex. However, due to the lack of an obvious subsite at the non reducing side, an alternate protocol has been chosen that incorporates bioc hemical information on the enzyme and features on the substrate shape as we ll. As a net result, ten subsites have been located consistent with the dis tribution of Ajandouz et al. (E. H. Ajandouz, J. Abe, B. Svensson, and G. M archis-Mouren, Biochimica Biophysica Acta, 1992, Vol. 1159, pp. 193-202) an d corresponding binding energies were estimated. Among them, two extreme su bsites (-6) and (+4), with stacking residues Y104 and Y211, respectively, h ave strong affinities with glucose rings added to the substrate. No other d eformation has been found for the new glucose rings added to the substrate; therefore, only ring A of the DP 10 fragment has a flexible form when inte racting with the inner stacking residues Y51. Global conservation of the he lical shape of the substrate can be postulated in spite of its significant distortion at subsite (-1). (C) 1999 John Wiley & Sons, Inc.