IDENTIFICATION AND PURIFICATION OF A STRESS ASSOCIATED NUCLEAR CARBOHYDRATE-BINDING PROTEIN (M(R)-33000) FROM RAT-LIVER BY APPLICATION OF ANEW PHOTOREACTIVE CARBOHYDRATE PROBE

A photoreactive a-D-glucose probe has been designed for the specific d etection of carbohydrate binding proteins (CBPs). The probe consists o f four parts: (i) an alpha-D-glucose moiety; (ii) the digoxigenin tag; (iii) the photoreactive cross-linker; and (iv) the lysyl-lysine backb one. After incubation with lectins in the dark, the probe is activated and cross-linked to the CBPs after being treated by several flashes. Using this method we have identified a new alpha-D-glucose CBP of M(r) = 33 000, termed CBP33, in the nuclei of rats exposed to transient im mobilization stress. Monoclonal antibodies were raised against the par tially purified protein and subsequently used to enrich CBP33. II was purified (>2400-fold) to apparent homogeneity from a 0.6 M nuclear sal t extract by two subsequent affinity chromatography steps (antibody-af finity as well as alpha-D-glucose affinity column).