The β-glucan svnthase from Lolium multiflorum: Detergent solubilization, purification using monoclonal antibodies, and photoaffinity labeling with a novel photoreactive pyrimidine analogue of uridine 5′-diphosphoglucose

Abstract

The membrane-bound β-glucan synthase from Italian ryegrass (Lolium multiflorum L.) endosperm cells has been solubilized by both non-ionic and zwitterionic detergents. A complex relationship exists between the ratio of (1→3)-, (1→4)-, and (1→3, 1→4)-β-glucan products of the solubilized enzyme, the cations present, and the concentration of the uridine 5′-diphosphoglucose substrate. Monoclonal antibodies directed against the β-glucan synthase complex were generated by immunization of mice with an unfractionated microsomal preparation. Hybridoma cell lines were screened using a combination of indirect enzyme-linked immunosorbent assay followed by an enzyme-capture assay. The purified monoclonal ..