Joshua S Sims

US Patent:

20110250613, Oct 13, 2011

Filed:

Jun 15, 2010

Appl. No.:

12/815740

Inventors:

Joshua J. Sims - Baltimore MD, US
Robert E. Cohen - Baltimore MD, US

Assignee:

THE JOHNS HOPKINS UNIVERSITY - Baltimore MD

International Classification:

G01N 33/566
C12N 9/00
C12N 1/21
C07K 14/00

US Classification:

435 71, 530350, 435183, 43525233, 436501

Abstract:

Linkage-specific polyubiquitin recognition is thought to make possible the diverse set of functional outcomes associated with ubiquitination. Thus far, mechanistic insight into this selectivity has been largely limited to single domains that preferentially bind to lysine 48-linked polyubiquitin (K48-polyUb) in isolation. A mechanism is proposed herein, linkage-specific avidity, in which multiple ubiquitin-binding domains are arranged in space so that simultaneous, high-affinity interactions are optimum with one polyUb linkage but unfavorable or impossible with other polyUb topologies and monoUb. The model used herein is human Rap80, which contains tandem ubiquitin interacting motifs (UIMs) that bind to K63-polyUb at DNA doublestrand breaks. The sequence between the Rap80 UIMs positions the domains for efficient avid binding across a single K63 linkage, thus defining selectivity. K48-specific avidity is also demonstrated in a different protein, ataxin-3. Using tandem UIMs, the general principles governing polyUb linkage selectivity and affinity in multivalent ubiquitin receptors are established.