Rabbit
antibodies directed against homogeneous
uridylate synthase multienzyme from mouse Ehrlich
ascites carcinoma precipitate both the orotidine-5'-monophosphate
decarboxylase (EC 4.1.1.23) and
orotate phosphoribosyltransferase (EC 2.4.2.10) activities of mouse and human erythrocyte
uridylate synthase. When the partially purified human
enzyme is used as
antigen the two activities coprecipitate with the same apparent titer; however, when the mouse
carcinoma protein was studied under the same conditions the
decarboxylase activity immunoprecipitated with significantly higher avidity than did the
transferase activity. Since the mouse multienzyme has been shown to be a single
polypeptide that contains both activities (McClard, R.W., Black, M.J., Livingstone, L.R. and Jones, M.E. (1980) Biochemistry 19, 4699-4706), these results were, at face value, surprising. However, when the mouse
orotate phosphoribosyltransferase activity (which is largely lost upon dilution into the immunoassay medium) was stabilized with 5-phosphoribosyl 1-pyrophosphate, both
enzyme activities displayed the same apparent antibody titer. The immunochemical studies indicate that the
antibodies, as a population, preferentially bind to a form or forms of the
enzyme which contain(s) denatured
transferase domains. A calculation based on a simple model yields a value of approximately 100 for the relative selectivity of the antibody for the denatured form of
uridylate synthase. These results illustrate an ambiguity that is inherent in the interpretation of immunochemical studies on such multienzymic
proteins; that is, it is possible to conclude incorrectly that two
enzyme activities are not functionally associated if one of the catalytic domains is particularly unstable and thereby displays greater immunoreactivity for the specific antiserum.