CRYSTAL AND MOLECULAR-STRUCTURE AT 0.16-NM RESOLUTION OF THE HYBRID BACILLUS ENDO-1,3-1,4-BETA-D-GLUCAN 4-GLUCANOHYDROLASE H(A16-M)

H(A16-M) is a hybrid endo-1,3-1,4-beta-D-glucan 4-glucanohydrolase fro m Bacillus. Its crystal structure was refined using synchrotron X-ray diffraction data up to a maximal resolution of 0.16 nm. The R value of the resulting model is 14.3% against 21 032 reflections > 2 sigma. 93 % of the amino acid residues are in the most favorable regions of the Ramachandran diagram, and geometrical parameters are in accordance wit h other proteins solved at high resolution. As shown earlier [Keitel, T., Simon, O., Borriss, R. & Heinemann, U. (1993) Proc. Natl Acad. Sci . USA 90, 5287-5291], the protein folds into a compact jellyroll-type beta-sheet structure. A systematic analysis of the secondary structure reveals the presence of two major antiparallel beta-sheets and a thre e-stranded minor mixed sheet. Amino acid residues involved in catalysi s and substrate binding are located inside a deep channel spanning the surface of the protein. To investigate the stereochemical cause of th e observed specificity of endo-1,3-1,4-beta-D-glucan 4-glucanohydrolas es towards beta-1,4 glycosyl bonds adjacent to beta-1,3 bonds, the hig h-resolution crystal structure has been used to model an enzyme-substr ate complex. It is proposed that productive substrate binding to the s ubsites p1, p2 and p3 of H(A16-M) requires a beta-1,3 linkage between glucose units bound to p1 and p2.