Genetic and biochemical analysis of alkaline phosphatase from inbred rat strains

摘要

Alkaline phosphatase (AP) was analyzed utilizing electrophoretic techniques and biochemical assays to establish genetic phenotypes and inheritance patterns for the purpose of genetic monitoring of inbred rats. The AP from rat kidney was further tested for functionality differences between the enzyme phenotypes.;Cellulose acetate electrophoresis (CAE) revealed that the AP of kidney, liver and small intestine contained three genetic phenotypes distinguished by their migration rates: AA (fast), BB (null), CC (slow). The phenotypes displayed a Mendelian inheritance pattern that was indicative of a single locus with multiple alleles. The AA and CC phenotypes were dominant over the BB phenotype whereas the AA and CC were codominant to each other. The phenotypic variation observed in CAE was due to differences in glycosylation of the enzyme.;Kidney AP from selected rat strains was purified and tested to determine if the different phenotypes identified created functional differences in the enzyme. The F$sb1$ hybrids displayed a single gene overdominance in the V$sb{rm max}$/K$sb{rm m}$ ratio over the inbred strains when either pH or temperature was varied. Other than overdominance no other functional differences were found between the inbreds animals and the F$sb1$ hybrids using the kinetic parameters in this study. The average Michaelis constant value was 1.43 mM using $rho$-nitrophenyl phosphate as the substrate. The average I$sb{50}$ for the inhibitors, L-homoarginine and imidazole, were 2.06 mM and 8.07 mM respectively.;This study demonstrated that the AP enzyme would be a useful biochemical enzyme marker for the genetic monitoring of inbred rats. It also demonstrated that a special case of heterosis was responsible for the difference between the inbred and F$sb1$ hybrids. Finally, an unusual small intestinal AP variant was found which may prove to be useful as an animal model system for human AP disease in that it resembled the human Kasahara AP variant.