Protein functional cycle viewed at atomic resolution: Conformational change and mobility in nitrophorin 4 as a function of pH and NO binding

Dmitry A. Kondrashov, Sue A. Roberts, Andrzej Weichsel, William R. Montfort

Research output: Contribution to journalArticle

54 Scopus citations

Abstract

The blood-sucking insect Rhodnius prolixus uses nitrophorin 4, a heme protein, to deliver nitric oxide (NO) to a victim, causing vasodilation and improved feeding. Binding of NO occurs at a ferric heme and is modulated by pH. NO binding at lower pH induces a large conformational change involving loops A-B and G-H that leads to distal pocket desolvation and protection of the nitrosyl heme complex. We have determined the crystal structures of Rhodnius nitrophorin 4 to ultrahigh resolution in four functional states: ±NO at pH = 7.4 and ±NO at pH = 5.6. The structure with NO at pH 7.4 (1.08 Å) is newly determined while the other complexes have been modeled to resolutions much greater than previously reported (1.0-0.85 Å). The ultrahigh resolution allowed us to resolve multiple conformers in binding-site loops, leading to a detailed description of the dynamics involved with storing NO in the insect salivary gland at low pH, and releasing NO in response to the increased pH of a victim's tissue. Strikingly, features for both the "open" and "closed" conformers exist under all conditions, suggesting that the flexible loops can transition with relative ease between conformational states. Yet, release of NO from rNP4 is much slower than found for other ferric heme proteins. The structures suggest that highly mobile loops can limit diffusion of diatomic molecules into and out of a protein cavity, a result with implications for the role of protein dynamics in function.

Original languageEnglish (US)
Pages (from-to)13637-13647
Number of pages11
JournalBiochemistry
Volume43
Issue number43
DOIs
StatePublished - Nov 2 2004

ASJC Scopus subject areas

  • Biochemistry

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