A rational protocol for the successful crystallization of l-amino-acid oxidase from Bothrops atrox


Autoria(s): ALVES, Raquel Melo; FELICIANO, Patricia Rosa; SAMPAIO, Suely Vilela; NONATO, Maria Cristina
Contribuinte(s)

UNIVERSIDADE DE SÃO PAULO

Data(s)

17/04/2012

17/04/2012

2011

Resumo

Despite the valuable contributions of robotics and high-throughput approaches to protein crystallization, the role of an experienced crystallographer in the evaluation and rationalization of a crystallization process is still crucial to obtaining crystals suitable for X-ray diffraction measurements. In this work, the difficult task of crystallizing the flavoenzyme l-amino-acid oxidase purified from Bothrops atrox snake venom was overcome by the development of a protocol that first required the identification of a non-amorphous precipitate as a promising crystallization condition followed by the implementation of a methodology that combined crystallization in the presence of oil and seeding techniques. Crystals were obtained and a complete data set was collected to 2.3 A resolution. The crystals belonged to space group P2(1), with unit-cell parameters a = 73.64, b = 123.92, c = 105.08 A, beta = 96.03 degrees. There were four protein subunits in the asymmetric unit, which gave a Matthews coefficient V (M) of 2.12 A3 Da-1, corresponding to 42% solvent content. The structure has been solved by molecular-replacement techniques.

FAPESP (Fundacao de Amparo a Pesquisa do Estado de Sao Paulo)[2005/54855-0]

FAPESP (Fundacao de Amparo a Pesquisa do Estado de Sao Paulo)[2007/51700-1]

FAPESP (Fundacao de Amparo a Pesquisa do Estado de Sao Paulo)[2009/10454-3]

Identificador

ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS, v.67, p.475-478, 2011

1744-3091

http://producao.usp.br/handle/BDPI/14883

10.1107/S1744309111003770

http://dx.doi.org/10.1107/S1744309111003770

Idioma(s)

eng

Publicador

WILEY-BLACKWELL

Relação

Acta Crystallographica Section F-structural Biology and Crystallization Communications

Direitos

closedAccess

Copyright WILEY-BLACKWELL

Palavras-Chave #seeding #crystallization in the presence of oil #l-amino-acid oxidases #snake venoms #Bothrops atrox #VIPER CALLOSELASMA-RHODOSTOMA #HUMAN PLATELET-AGGREGATION #SNAKE-VENOM #PROTEIN CRYSTALLIZATION #PURIFICATION #ANTIBACTERIAL #MECHANISM #LAAO
Tipo

article

original article

publishedVersion