Role of the intramolecular hydrogen bond network in the inhibitory power of chymotrypsin inhibitor 2

Evette S Radisky, Chia Jung Karen Lu, Gene Kwan, Daniel E. Koshland

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

A series of mutants of chymotrypsin inhibitor 2 (CI2), at residues involved in intramolecular interactions that shape and constrain the binding loop, were studied to determine their relative importance for inhibition of the serine protease subtilisin BPN′, and for resistance of the inhibitor to proteolysis. These functional properties were investigated in tandem with the crystal structures of the mutant inhibitor-enzyme complexes. A dense hydrogen bonding network that supports the binding loop in the vicinity of the scissile bond was found to be important both for enzyme affinity and for stability to proteolysis. Structural analysis, in combination with biochemical measurements, allows differentiation of the structural components most important for resistance to proteolysis and/or binding. The most critical participating residues in the network were found to be Thr-58, Glu-60, Arg-65, and Gly-83. Glu-60 is more important for resistance to proteolysis than for binding, while Arg-65 and two other Arg residues play a greater role in binding than in resistance to proteolysis. Structural comparisons reveal a wide variety of subtle conformational changes in response to mutation, with built-in robustness in the hydrogen bond network, such that loss of one contact is compensated by other new contacts.

Original languageEnglish (US)
Pages (from-to)6823-6830
Number of pages8
JournalBiochemistry
Volume44
Issue number18
DOIs
StatePublished - May 10 2005
Externally publishedYes

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Proteolysis
Hydrogen
Hydrogen bonds
Subtilisins
Enzyme Stability
Serine Proteases
Enzyme Inhibitors
Hydrogen Bonding
Structural analysis
Crystal structure
chymotrypsin inhibitor 2
Mutation
Enzymes

ASJC Scopus subject areas

  • Biochemistry

Cite this

Role of the intramolecular hydrogen bond network in the inhibitory power of chymotrypsin inhibitor 2. / Radisky, Evette S; Lu, Chia Jung Karen; Kwan, Gene; Koshland, Daniel E.

In: Biochemistry, Vol. 44, No. 18, 10.05.2005, p. 6823-6830.

Research output: Contribution to journalArticle

Radisky, Evette S ; Lu, Chia Jung Karen ; Kwan, Gene ; Koshland, Daniel E. / Role of the intramolecular hydrogen bond network in the inhibitory power of chymotrypsin inhibitor 2. In: Biochemistry. 2005 ; Vol. 44, No. 18. pp. 6823-6830.
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