Specificity in Protein-Protein Recognition: Conserved Im9 Residues Are the Major Determinants of Stability in the Colicin E9 DNase-Im9 Complex
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- 作者：Russell Wallis ; Kit-Yi Leung ; Michael J. Osborne ; Richard James ; Geoffrey R. Moore ; and Colin Kleanthous
- 刊名：Biochemistry
- 出版年：1998
- 出版时间：January 13, 1998
- 年：1998
- 卷：37
- 期：2
- 页码：476 - 485
- 全文大小：251K
- 年卷期：v.37,no.2(January 13, 1998)
- ISSN：1520-4995
文摘
The endonuclease group of E colicins are a family of bacterialtoxins whose cytotoxic activityin a producing host is inactivated by a specific immunity protein.The DNase of colicin E9 can be boundand inhibited by both cognate and noncognate immunity proteins, thedissociation constants for whichspan a range of 12-orders of magnitude. DNase binding specificityof the immunity proteins is governedprimarily by helix II, the sequence of which is variable in this familyof proteins. Heteronuclear NMRexperiments have identified helix III along with helix II as the likelyDNase binding site, although otherregions of Im9 also showed perturbations on binding the E9 DNase.In the present work, we have usedthe NMR experiments as a guide for alanine scanning mutagenesis of Im9.Our data show that helicesII and III of Im9 are indeed the DNase binding site and in additionquantitate the relative binding energyassociated with each helix. We find that the conserved residues ofhelix III make the largest relativecontribution toward E9 DNase binding. In conjunction with previousstudies, the data suggest thatspecificity in the colicin-immunity system is governed by a dualrecognition mechanism in which highlystabilizing interactions emanating from the conserved regions of animmunity protein act as the bindingsite anchor and these are modulated by interactions from neighboring,nonconserved amino acid residues.This modulation is likely to take the form of both favorable andunfavorable interactions, the balance ofwhich define the specificity of the protein-protein interaction.The generality of such a dual recognitionmechanism in other systems is also discussed.