DNA repair proteins participate in extensive proteinÀ protein interactions that promote the formation of DNA repair complexes. To understand how complex formation affects protein function during base excision repair, we used SpyCatcher/ SpyTag ligation to produce a covalent complex between human uracil DNA glycosylase (UNG2) and replication protein A (RPA). Our covalent "RPAÀ SpyÀ UNG2" complex could identify and excise uracil bases in duplex areas next to ssDNAÀ dsDNA junctions slightly faster than the wild-type proteins, but this was highly dependent on DNA structure, as the turnover of the RPAÀ SpyÀ UNG2 complex slowed at DNA junctions where RPA tightly engaged long ssDNA sections. Conversely, the enzymes preferred uracil sites in ssDNA where RPA strongly enhanced uracil excision by UNG2 regardless of ssDNA length. Finally, RPA was found to promote UNG2 excision of two uracil sites positioned across a ssDNAÀ dsDNA junction, and dissociation of UNG2 from RPA enhanced this process. Our approach of ligating together RPA and UNG2 to reveal how complex formation affects enzyme function could be applied to examine other assemblies of DNA repair proteins.

No abstract

Sargatal, J. (1992Sargatal, J. ( -1997): Handbook of the birds of the world. Volumes 1 4 . Lynx Edicions, Kassai, T.; Corder0 del Campillo, M.; Euzeby, J.; Gaafar, S.; Hiepe, T.; Himonas, C. A. (1988): Standardized Kirchner, J. V. (1992): Diccionario multilingtie de especias marinas para el mundo hispano. Ministerio de Mack, R.; Meissonnier, E. (199 1): Dictionnaire des tennes v6t6rinaires et animaliers, franpk-anglais, anglais-Mitchell-Jones, A. J.; Amori, G.; Bogdaiiowicz, W.; KryStufek, B. et al. (1999): The atlas of European mammals. Murphy, F. A.; Gibbs, E. P. J.; Honinek. M. C.; Studdert, M. J. (1999): Veterinary virology. 3rd edition. Academic National Library of Medicine, USA. Medical subject headings [current issue]. Nickel, R.; Schummer, A.; Seiferle, E. (Begriinder) (1960-1999): Lehrbuch der Anatomie der Haustiere.

No abstract