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Escherichia coli Expression, Purification, Crystallization, and Structure Determination of Bacterial Cohesin–Dockerin Complexes

2012; Academic Press; Linguagem: Inglês

10.1016/b978-0-12-415931-0.00021-5

1557-7988

Joana L. A. Brás, Ana Luı́sa Carvalho, Aldino Viegas, Shabir Najmudin, Victor D. Alves, José A. M. Prates, L.M.A. Ferreira, Maria João Romão, Harry J. Gilbert, C.M.G.A. Fontes,

Enzyme Production and Characterization

Cellulosomes are highly efficient nanomachines that play a fundamental role during the anaerobic deconstruction of complex plant cell wall carbohydrates. The assembly of these complex nanomachines results from the very tight binding of repetitive cohesin modules, located in a noncatalytic molecular scaffold, and dockerin domains located at the C-terminus of the enzyme components of the cellulosome. The number of enzymes found in a cellulosome varies but may reach more than 100 catalytic subunits if cellulosomes are further organized in polycellulosomes, through a second type of cohesin-dockerin interaction. Structural studies have revealed how the cohesin-dockerin interaction mediates cellulosome assembly and cell-surface attachment, while retaining the flexibility required to potentiate catalytic synergy within the complex. Methods that might be applied for the production, purification, and structure determination of cohesin-dockerin complexes are described here.