Developmental expression of the embryonic chicken brain DNA polymerase alpha and its binding with monoclonal antibodies against human KB cell DNA polymerase alpha.
Changes in DNA polymerase alpha activity accompanying tissue development have been well established in several systems. In most cases, DNA polymerase alpha activity decreases with development. Here, we report observed changes in DNA polymerase alpha activity throughout embryonic chicken brain (ECB) development. The level of DNA polymerase alpha activity was found to gradually decrease by 60% (2.3 to 0.8 nmol of [3H]dCMP incorporated/mg protein/h) between 9- and 19-day-old ECB. An enzyme-linked immunosorbent assay of DNA polymerase alpha utilizing monoclonal antibody SJK 237-71 (human KB cell DNA pol-alpha binder) also demonstrated a gradual decrease (up to 60%) of antigen over this same range of development. Analysis of DNA polymerase alpha from 11- and 19-day-old ECB by a 10 to 30% glycerol density gradient revealed a high molecular weight peak sedimenting near catalase (11.3 S) with activity at the 11th day being approximately 3-fold greater than activity at the 19th day. A Western immunoblot analysis utilizing monoclonal antibody SJK 237-71 (against human KB cell DNA polymerase alpha) showed a decrease in DNA polymerase alpha from 186 kilodaltons in 9- and 11-day ECB cell-free extracts to 120 kilodaltons in extracts from 13- to 19-day ECB. The conversion of DNA polymerase alpha from a higher to a lower molecular weight form may be a regulatory mechanism in eukaryotic DNA replication.
Ray
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Department of Chemistry and Biochemistry, University of Notre Dame, Indiana 46556.
Kelley
T J
TJ
Campion
S
S
Seve
A P
AP
Basu
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001921:Brain / Q000196:embryology / Q000201:enzymology; D002642:Chick Embryo; D003429:Cross Reactions; D004257:DNA Polymerase II / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification; D019915:DNA Primase; D006801:Humans; D007624:KB Cells / Q000201:enzymology; D012316:RNA Nucleotidyltransferases / Q000302:isolation & purification; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D019915:DNA Primase; D012316:RNA Nucleotidyltransferases; D004257:DNA Polymerase II