ALPHA-1,4-D-GLUCAN PHOSPHORYLASE OF GRAM-POSITIVE CORYNEBACTERIUM-CALLUNAE - ISOLATION, BIOCHEMICAL-PROPERTIES AND MOLECULAR SHAPE OF THE ENZYME FROM SOLUTION X-RAY-SCATTERING

The alpha-1,4-D-glucan phosphorylase from gram-positive Corynebacteriu m callunae has been isolated and characterized, The enzyme is inducibl e approx, 2-fold by maltose, but remarkably not repressed by D-glucose , The phosphorylase is a homodimer with a stoichiometric content of th e cofactor pyridoxal 5'-phosphate per 88-kDa protein subunit, The spec ificity constants (k(cat)/K-m,K-glucan) in the directions of glucan sy nthesis and degradation are used for the classification of the enzyme as the first bacterial starch phosphorylase. A preference for large ov er small substrates is determined by variations in the apparent bindin g constants rather than catalytic-centre activities. The contribution of substrate chain length to binding energy is explained assuming two glucan binding sites in C, callunae phosphorylase: an oligosaccharide binding site composed of five subsites and a high-affinity polysacchar ide site separated from the active site. A structural model of the mol ecular shape of the phosphorylase was obtained from small-angle soluti on X-ray scattering measurements. A flat, slightly elongated, ellipsoi dal model with the three axes related to each other as 1 :(0.87-0.95): 0.43 showed scattering equivalence with the enzyme molecule. The model of C, callunae phosphorylase differs from the structurally well-chara cterized rabbit-muscle phosphorylase in size and axial dimensions.