Entamoeba histolyticaEhGEF1 structure and mutational analysis: New specific residues critical for function.
This paper reports the EhGEF1-EhRacG and EhGEF1-EhRho1 molecular complexes from Entamoeba histolytica. The not conserved amino acids Gln201,Tyr299, Gln302, Lys312, Asn313, Phe314 and Ile324 were localized, by means of an in silico computational analysis, at the interface of the exposed face from the DH domain of EhGEF1, which are important to establish the contact with its target GTPases EhRacG and EhRho1. Functional studies of nucleotide exchange of Phe314Ala mutant showed a decrement of 80% on EhRacG GTPase; in contrast the Ile324Ala mutant exhibited a reduction of 77%, specifically on EhRho1; meanwhile the Gln302Ala mutant showed a reduction of approximately 50% on the exchange activity for both GTPases. Moreover, the functional studies of the protein EhGEF1 mutants in the conserved residues Thr194Ala, Asn366Ala and Glu367Ala indicated that contrary to what has been reported for other systems, the mutation of these residues did not alter considerably its catalytic activity.
Hernández-Cuevas
N A
NA
Departamento de Biomedicina Molecular, Centro de Investigación y de Estudios Avanzados, México, D.F., Mexico.
Campos-Parra
A D
AD
Almaraz-Barrera
Ma de J
Mde J
Aguilar-Rojas
A
A
González-de la Rosa
C H
CH
Sosa-Peinado
A
A
Hernández-Rivas
R
R
Rojo-Domínguez
A
A
Vargas
M
M
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D018118:Chloride Channels / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004252:DNA Mutational Analysis; D004748:Entamoeba histolytica / Q000235:genetics / Q000502:physiology*; D019204:GTP-Binding Proteins / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D011485:Protein Binding; D025941:Protein Interaction Mapping; D020836:Protein Structure, Quaternary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
D018118:Chloride Channels; D015800:Protozoan Proteins; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins