A one-residue switch reverses the orientation of a heme b cofactor. Investigations of the ferriheme NO transporters nitrophorin 2 and 7 from the blood-feeding insect Rhodnius prolixus.
Biochemistry
; 48(2): 235-41, 2009 Jan 20.
Article
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| MEDLINE
| ID: mdl-19140692
This study represents the identification of a single amino acid residue that has the major responsibility for the isomeric orientation of a heme b cofactor in a ferriheme protein. The insertion of hemin b into the asymmetric environment of a protein pocket facilitates two cofactor orientations, A and B, which is often called "heme rotational disorder". The proteins studied herein are nitrophorins, a class of ferriheme proteins found in the saliva of the blood-sucking insect Rhodnius prolixus, in this case NP2 and NP7. NMR spectroscopy (pH* 5.5) of the imidazole complex of NP7 revealed solely the A orientation, whereas NP2 shows primarily the B orientation ( approximately 1:5 A:B). The glutamate 27 residue in NP7 is an obvious difference in the heme pocket compared to those of NP1-4, all of which present a valine residue [valine 24 (NP2 and NP3) or valine 25 (NP1 and NP4)] at the same position. Consequently, the mutant NP2(V24E) was prepared and shown to reverse the heme orientation to exclusively A, whereas NP7(E27V) revealed an approximately 1:3 A:B ratio. The reversal A <--> B following the change glutamine <--> valine was further indicated in circular dichroism (CD) spectroscopy with a positive (A) or negative (B) Deltaepsilon of the heme Soret band. Moreover, CD spectroscopy was applied to the mutant NP7(E27Q) and indicated mainly the A orientation, which allows us to conclude that the steric hindrance provided by the glutamate residue is responsible for the heme orientation rather then the carboxylate charge.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Rhodnius
/
Proteínas y Péptidos Salivales
/
Proteínas de Insectos
/
Hemo
/
Hemoproteínas
/
Hemina
Límite:
Animals
Idioma:
En
Revista:
Biochemistry
Año:
2009
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Estados Unidos