Specificity of Solubilized Yeast Glycosyl Transferases for Polyprenyl Derivatives

Abstract

Four glycosyl transferases solubilized by detergents from Saccharomyces cerevisiae membranes have been tested for their specificity towards various polyprenols differing in chain length and saturation. The formation of dolichyl diphosphate‐( N ‐acetylglucosamine) 1‐2 from uridine diphosphate N ‐acetylglucosamine and dolichyl monophosphate showed an obligatory requirement for α‐saturated polyprenols. The apparent affinity for α‐saturated polyprenyl phosphates improves with increasing chain length; those shorter than seven isoprene units were inactive. The rate of transfer of the chitobiosyl unit from dolichyl diphosphate chitobiose to the hexapeptide Tyr‐Asn‐Leu‐Thr‐Ser‐Val followed the sequence: α‐dihydropolyprenyl (C 55 ) >(C 80 > (C 100 ). The enzyme transferring the mannosyl group from guanosine diphosphate mannose to dolichyl monophosphate also showed a clear dependence on α‐saturation and chain length. In this case the chain length affects mainly the maximal velocity of the reaction. The highest value was obtained with C 55 α‐dihydropolyprenyl phosphate (100 nmol × h −1 × mg protein −1 ), whereas the value with C 35 α‐dihydropolyprenyl phosphate was only 8 nmol × h −1 × mg protein. −1 . The mannosyl transfer from dolichyl monophosphate mannose to dinitrophenylated tetrapeptide Asn‐Ala‐Thr‐Vol to form on O‐glycosidic bond strongly depended on α‐saturation and chain length of the polyprenyl phosphate. The rate of transfer correlated directly with the length of the polyprenyl residue. With C 80 α‐dihydropolyprenol less than half the rate of the C 100 compound was observed, whereas C 55 and C 35 α‐dihydropolyprenol are only 20 and 10% as active, respectively.