Factors Affecting Protein and Glycoprotein Synthesis by Rat Kidney Mitochondria <i>in vitro</i>

Abstract

Incorporation of ¹⁴C-labeled leucine and mannose were used as indices of protein and glycoprotein synthesis by isolated rat kidney mitochondria. Uninephrectomy, certain salts including NaCl and heavy metals, detergents such as digitonin and lysophosphatidylcholine, and various antibiotics were studied for their effect in this in vitro assay system. Amino acid incorporation by mitochondria into macromolecules was adversely affected by a number of the conditions studied while carbohydrate incorporation was less frequently influenced. The number of variables capable of influencing the assay system indicate that caution must be used in the interpretation of in vitro enzyme reactions especially in the study of experimental nephropathies. Alterations in mitochondrial macromolecular synthesis may be a feature of various types of renal injury though the importance of mitochondrial biosynthetic activity to the overall cell response remains to be established.

Alterations of Mitochondrial Properties in Folate Nephropathy

“…The absence of changes in the activities of succinic dehydrogenase and monoamine oxidase suggests a rather subtle and selective disturbance of amino acid incorpo ration. Previous studies indicated that amino acid in corporation in vitro is sensitive to the concentrations of NaCl and CaCle in the medium [12]. With tissue injury impairing the ability of the membrane-active transport systems to preserve normal electrolyte conditions, intra cellular and intramitochondrial Na and Ca concentra tions may rise [5.9,13].…”

“…The incorporation of ' 'C-leucine by mitocondria in vitro was measured as previously described [12], Assays were performed in quadruplicate and correct for radioactivity incorporated in the pres ence of chloramphenicol, a known inhibitor of mitochondrial protein synthesis. Results are expressed as cpm/mg protein.…”

“…The incorporation of "C-mannose from GDP-"C-mannose into pellets washed with 10% trichloroacetic acid and ethanol-ether (2 :1V/V) was quantitated as previously detailed [12], Results are as cpm/mg protein.…”