Organ specificity of glucocorticoid-sensitive tyrosine aminotransferase. Separation from aspartate aminotransferase isoenzymes.
In order to study whether hormone-sensitive tyrosine aminotransferase exists in tissues other than liver, we have devised means to separate the liver-specific enzyme from other enzymes that transaminate tyrosine and to distinguish between the authentic enzyme and the principal "pseudotyrosine aminotransferases," which are the isoenzymes of aspartate aminotransferase. We accomplish this by suppressing proteolysis of the authentic enzyme using a buffer of pH 8.0 containing 0.1 M potassium chloride; enzyme extracted from liver in this buffer migrates as a single peak during chromatography on hydroxylapatite and represents the undegraded native form. A much smaller peak of tyrosine aminotransferase activity elutes at higher ionic strength and corresponds to a mixture of mitochondrial aspartate aminotransferase and partially degraded tyrosine aminotransferase. Cytosolic aspartate aminotransferase, in contrast, adsorbs weakly to the hydroxylapatite column and transaminates tyrosine very poorly although it readily utilizes monoiodotyrosine. The aspartate aminotransferase isoenzymes separate completely from tyrosine aminotransferase during chromatography on DEAE-Sepharose CL-6B. By combining these techniques with the use of specific antibodies, we show that brain, heart, and kidney do not contain tyrosine aminotransferase. Furthermore, we locate both isoenzymes of aspartate aminotransferase on polyacrylamide gels and show that both react histochemically as tyrosine aminotransferases when monoiodotyrosine is used as substrate. Use of these techniques, therefore, permits unambiguous identification of tyrosine aminotransferase and its separation from the background of nonspecific transamination.
D000818:Animals; D001219:Aspartate Aminotransferases / Q000302:isolation & purification*; D002852:Chromatography, Ion Exchange; D005260:Female; D005938:Glucocorticoids / Q000494:pharmacology*; D007527:Isoenzymes / Q000302:isolation & purification*; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008297:Male; D009206:Myocardium / Q000201:enzymology; D051381:Rats; D011919:Rats, Inbred Strains; D014018:Tissue Distribution; D014444:Tyrosine Transaminase / Q000302:isolation & purification*
D005938:Glucocorticoids; D007527:Isoenzymes; D001219:Aspartate Aminotransferases; D014444:Tyrosine Transaminase