Ultraviolet light-induced crosslinking of HnRNA to informofer proteins in vitro

RNA-protein interaction in the 30S subunits of rat liver hnRNP has been studied by crosslinking of informofer proteins to hnRNA induced by UV irradiation. Irradiation of 30S particles with 254 nm UV light in doses of 1 1 x 10(5) erg/mm2 leads to the extensive crosslinking hnRNA to informofer proteins. The crosslinked material was analyzed either by resedimentation in a 15-30% sucrose gradient in the presence of 3 M guanidine-HCl and 1 M NaCl or by centrifugation in a Cs2SO4 density gradient containing guanidine-HCl and sarkosyl. The crosslinked complexes sedimented at about 25S in the sucrose gradient and proved to be heterogeneous in isopycnic centrifugation experiments. The proteins of the crosslinked complexes were analyzed by polyacrylamide gel electrophoresis. Proteins with Mr values of 70 000, 58 000, 43 000 and 40 000 appeared to be crosslinked with hnRNAs of the 30S particles. In the unirradiated 30S particles after centrifugation in the CS2SO4 density gradient containing guanidine-HCl and sarkosyl two minor proteins were observed with Mr values of 70 000 and 58 000, banded in density zones characteristic for free RNA.