Procedures are being developed to differentially label with N-etylmaleimide or fluorodinitrobenzene the transporters which constitute the L-, Gly- and ASC systems for neutral amino acids and the systems of hexoses and nucleosides in the bovine lens. Both 3H- and 14C-radioisotopes of the group reagents are to be utilized. The differential labeling will be detected by counting sections of polyacrylamide gels after electrophoresis in sodium dodecyl sulfate. Labeled plasma membranes (PM) will be solubilized in Triton X-100, octylglucoside or zwitterionicsulfobetaines, and the transporters purified further by fractional precipitation or column chromatography, identifying pooled differentially labeled material by the 3H/14C ratio. PM are to be prepared from the bovine lens by a procedure which involves simultaneous extraction with 7 M guanidine hydrochloride and citraconylation (Roy et al, Exp. Eye Res. 28,353,1979). Attempts will be made to detect the differential labeling in cortical and epithelial homogenates and in PM prepared from both epithelium and cortex. Binding studies are also to be done on various fractions mentioned above. Analysis of transport of Ca ions in the bovine lens will be continued, and studies initiated in the toad's lens in collaboration with Dr. O. Candia. Effects will be examined of ruthenium red and La ions, inhibitors of (Ca ions plus Mg ions) -activated ATPase, on potential, resistivity, fluxes of Ca ions (45Ca) and Na ion, Ca ions -exchange. Additional experiments will be performed to characterize transport of CA ions and organic nutrients in the human lens.
NIH ExPorter
NIH ExPorter