Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of AHP2, a signal transmitter protein from Arabidopsis thaliana.
Acta Crystallogr Sect F Struct Biol Cryst Commun
; 69(Pt 2): 158-61, 2013 Feb 01.
Article
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| MEDLINE
| ID: mdl-23385758
ABSTRACT
Histidine-containing phosphotransfer proteins from Arabidopsis thaliana (AHP1-5) act as intermediates between sensor histidine kinases and response regulators in a signalling system called multi-step phosphorelay (MSP). AHP proteins mediate and potentially integrate various MSP-based signalling pathways (e.g. cytokinin or osmosensing). However, structural information about AHP proteins and their importance in MSP signalling is still lacking. To obtain a deeper insight into the structural basis of AHP-mediated signal transduction, the three-dimensional structure of AHP2 was determined. The AHP2 coding sequence was cloned into pRSET B expression vector, enabling production of AHP2 fused to an N-terminal His tag. AHP2 was expressed in soluble form in Escherichia coli strain BL21 (DE3) pLysS and then purified to homogeneity using metal chelate affinity chromatography and anion-exchange chromatography under reducing conditions. Successful crystallization in a buffer which was optimized for thermal stability yielded crystals that diffracted to 2.5 Å resolution.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Fosfotransferasas
/
Transducción de Señal
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Arabidopsis
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Proteínas de Arabidopsis
Idioma:
En
Revista:
Acta Crystallogr Sect F Struct Biol Cryst Commun
Año:
2013
Tipo del documento:
Article
País de afiliación:
República Checa