PRECUSOR FORMS OF RAT SMALL INTESTINAL LACTASE

Abstract

High molecular weight precursor forms of various intestinal brush border enzymes have been demonstrated by pulse-labelling of intestinal explants maintained in culture. In the present study we have examined in vivo precursor forms of rat intestinal lactase. Fasted infant rats were injected (IP) with [35S]-methionine and killed after 0.5, 2, 4, and 18 H. The short fast was necessary to maximize incorporation of label. Lactase forms were isolated from Triton X-100 extracts of the intestinal homogenates by immunoprecipitation techniques using highly selective antisera. The isolated enzyme forms were separated by SDS-electrophoresis and examined by fluorography. Labelling within 30 min occurred in precursor forms of apparent mol. wts of 170, 125, and 90 Kd. A further precursor form of 200 Kd appeared within 2 H. All labelled precursor forms were no longer evident by 4 H. Apparent labelled brush border forms of lactase of 220, 130, and 95 Kd were evident by 2 H and were still present after 18 H. Using similar methodology, we have examined precursors of sucrase-isomaltase, with confirmation of existence of mature and immature precursor forms of 250 and 240 Kd respectively. Demonstration of lactase precursor forms determined in vivo provides confirmation of similar in vitro findings. These studies are consistent with the initial intra-cellular synthesis of high molecular weight precursor forms of lactase with subsequent transfer to brush border sites.