1K5R

YAP65 WW domain S24-Amino-Ethylsulfanyl-Acetic Acid mutant

Summary for 1K5R

• Entry DOI: 10.2210/pdb1k5r/pdb
• Related: 1EG3 1EG4 1EOM 1I5H 1JMQ
• Descriptor: 65 KDA YES-ASSOCIATED PROTEIN, Fragment of WBP-1 (2 entities in total)
• Functional Keywords: ww domain, yap65, beta-sheet proteins, stability of beta sheets, signaling protein
• Cellular location: Cytoplasm: P46937
• Total number of polymer chains: 2
• Total formula weight: 5695.36

Authors

Ferguson, N.,Pires, J.R.,Toepert, F.,Johnson, C.M.,Pan, Y.P.,Volkmer-Engert, R.,Schneider-Mergener, J.,Daggett, V.,Oschkinat, H.,Fersht, A.R. (deposition date: 2001-10-12, release date: 2001-11-02, Last modification date: 2024-10-30)

Primary citation

Ferguson, N.,Pires, J.R.,Toepert, F.,Johnson, C.M.,Pan, Y.P.,Volkmer-Engert, R.,Schneider-Mergener, J.,Daggett, V.,Oschkinat, H.,Fersht, A.
Using flexible loop mimetics to extend phi-value analysis to secondary structure interactions.
Proc.Natl.Acad.Sci.USA, 98:13008-13013, 2001

PubMed Abstract: Chemical synthesis allows the incorporation of nonnatural amino acids into proteins that may provide previously untried probes of their folding pathway and thermodynamic stability. We have used a flexible thioether linker as a loop mimetic in the human yes kinase-associated protein (YAP 65) WW domain, a three-stranded, 44-residue, beta-sheet protein. This linkage avoids problems of incorporating sequences that constrain loops to the extent that they significantly change the nature of the denatured state with concomitant effects on the folding kinetics. An NMR solution structure shows that the thioether linker had little effect on the global fold of the domain, although the loop is apparently more dynamic. The thioether variants are destabilized by up to 1.4 kcal/mol (1 cal = 4.18 J). Preliminary Phi-value analysis showed that the first loop is highly structured in the folding transition state, and the second loop is essentially unstructured. These data are consistent with results from simulated unfolding and detailed protein-engineering studies of structurally homologous WW domains. Previously, Phi-value analysis was limited to studying side-chain interactions. The linkers used here extend the protein engineering method directly to secondary-structure interactions.

PubMed: 11687614
DOI: 10.1073/pnas.221467398

Experimental method

SOLUTION NMR