Mechanistic insights revealed by a UBE2A mutation linked to intellectual disability

Agradecimentos: LNBio/CNPEM; Brazilian National Council for Scientific and Technological Development (CNPq)National Council for Scientific and Technological Development (CNPq) [306879/2014-0, 310536/2014-6, 422790/2016-8]; Sao Paulo Research Foundation (FAPESP)Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP) [2012/50981-5, 2013/08028-1, 2015/06281-7]; NIH/NIGMSUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Institute of General Medical Sciences (NIGMS) [R01 GM088055] Ubiquitin-conjugating enzymes (E2) enable protein ubiquitination by conjugating ubiquitin to their catalytic cysteine for subsequent transfer to a target lysine side chain. Deprotonation of the incoming lysine enables its nucleophilicity, but determinants of lysine activation remain poorly understood. We report a novel pathogenic mutation in the E2 UBE2A, identified in two brothers with mild intellectual disability. The pathogenic Q93E mutation yields UBE2A with impaired aminolysis activity but no loss of the ability to be conjugated with ubiquitin. Importantly, the low intrinsic reactivity of UBE2A Q93E was not overcome by a cognate ubiquitin E3 ligase, RAD18, with the UBE2A target PCNA. However, UBE2A Q93E was reactive at high pH or with a lowpK a amine as the nucleophile, thus providing the first evidence of reversion of a defective UBE2A mutation. We propose that Q93E substitution perturbs the UBE2A catalytic microenvironment essential for lysine deprotonation during ubiquitin transfer, thus generating an enzyme that is disabled but not dead FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQ Fechado