60th residues of ubiquitin and Nedd8 are located out of E2-binding surfaces, but are important for K48 ubiquitin-linkage

Choi, Yun-Seok; Jeon, Young Ho; Ryu, Kyoung-Seok; Cheong, Chaejoon

doi:10.1016/j.febslet.2009.09.034

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Volume 583, Issue 20 , Pages 3323-3328, 20 October 2009

60th residues of ubiquitin and Nedd8 are located out of E2-binding surfaces, but are important for K48 ubiquitin-linkage

Edited by Noboru Mizushima

Division of Magnetic Resonance, Korea Basic Science Institute Ochang Campus, Cheongwon-Gun, Ochang-Eup, Yangcheong-Ri 804-1, Chungcheongbuk-Do 363-883, South Korea

Department of Bio-Analytical Science, University of Science and Technology, Daejon 305-333, South Korea

Received 3 August 2009; received in revised form 13 September 2009; accepted 16 September 2009. published online 24 September 2009.

Abstract

Nedd8, a ubiquitin-like modifier, is covalently attached to various proteins. Although Nedd8 has higher sequence identity (57%) with ubiquitin, its conserved K48 residue cannot form covalent linkage with ubiquitin. To decipher the reason why Nedd8 cannot be an effective ubiquitin-acceptor, we compared the non-covalent interaction between Nedd8 and ubiquitin for various E2s using cross-saturation NMR technique. However, both Nedd8 and ubiquitin displayed almost identical non-covalent E2-binding properties. The K60 of Nedd8 was not present at the E2-binding surface, but its mutation to Asn converted Nedd8 into a ubiquitin-acceptor. The N60 ubiquitin mutants also displayed a decreased ubiquitin-accepting activity. These results suggest the presence of an uncharacterized determinant for the K48 ubiquitin-linkage that is not related to non-covalent E2-bindings.