Castaneda CA, Dixon EK, Walker O, Chaturvedi A, Nakasone MA, Curtis JE, Reed MR, Krueger S, Cropp TA, Fushman D

Polyubiquitination, a critical protein post-translational modification, signals for a diverse set of cellular events via the different isopeptide linkages formed between the C terminus of one ubiquitin (Ub) and the É›-amine of K6, K11, K27, K29, K33, K48, or K63 of a second Ub. We assembled di-ubiquitins (Ub2) comprising every ...

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lysine linkage and examined them biochemically and structurally. Of these, K27-Ub2 is unique as it is not cleaved by most deubiquitinases. As this remains the only structurally uncharacterized lysine linkage, we comprehensively examined the structures and dynamics of K27-Ub2 using nuclear magnetic resonance, small-angle neutron scattering, and in silico ensemble modeling. Our structural data provide insights into the functional properties of K27-Ub2, in particular that K27-Ub2 may be specifically recognized by K48-selective receptor UBA2 domain from proteasomal shuttle protein hHR23a. Binding studies and mutagenesis confirmed this prediction, further highlighting structural/recognition versatility of polyubiquitins and the potential power of determining function from elucidation of conformational ensembles.

Date: Mar. 01, 2016

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