Monoclonal antimyogenin antibodies define epitopes outside the bHLH domain where binding interferes with protein-protein and protein-DNA interactions.
We have developed a panel of monoclonal antibodies against rat myogenin, a skeletal muscle regulatory protein of the bHLH family. Some of these monoclonals have been widely used by others, and details of their production are presented. Mapping the epitopes by immunoprecipitation of myogenin deletion mutants demonstrates that this panel recognizes epitopes spanning the entire molecule outside the HLH region. Four antibodies against epitopes outside the bHLH region interfere with the binding of myogenin/E-protein heterodimers to DNA sequences containing the myogenin heterodimer consensus recognition site. Three of these epitopes are partially masked in the heterodimers; antibody binding to these epitopes reduces the interactions between myogenin and E12. This suggests that surfaces outside the HLH dimerization domain may contribute to the stability of myogenin/E12 complexes. The binding of one antibody to its epitope did not appear to affect the myogenin/E12 interaction but nonetheless interfered with the binding of the complex to DNA, suggesting that this epitope lies near to a surface occupied by DNA.
University of Texas Southwestern Medical Center, Department of Cell Biology and Neuroscience, Dallas 75235, USA.
W E
WE
Wright
I
I
Dac-Korytko
K
K
Farmer
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000494:pharmacology; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D000939:Epitopes / Q000276:immunology*; D018257:Helix-Loop-Helix Motifs; D046911:Macromolecular Substances; D051379:Mice; D008807:Mice, Inbred BALB C; D018008:Myogenin / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation; D055503:Protein Multimerization; D051381:Rats; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors
D000911:Antibodies, Monoclonal; D004268:DNA-Binding Proteins; D000939:Epitopes; D046911:Macromolecular Substances; C503801:Myog protein, mouse; C503803:Myog protein, rat; D018008:Myogenin; D051820:TCF Transcription Factors; C544913:Tcf7l1 protein, mouse; C553060:Tcf7l1 protein, rat; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors; D004247:DNA