IscS and IscU, the two central protein components of the iron sulfur cluster assembly machinery, form a complex that is still relatively poorly characterized. In an attempt to standardize the purification of these proteins for structural studies we have developed a protocol to produce them individually in high concentration and purity. We show that IscS is a rather robust protein as long as it is produced in a PLP loaded form and that this co-factor is essential for fold stability and enzyme activity. In contrast to previous evidence, we also propose that, in contrast with previous evidence, IscU is a thermodynamically stable protein with a well defined fold but, when produced in isolation, is a 'complex-orphan protein' that is prone to unfolding if not stabilised by a co-factor or a protein partner. Our work will facilitate further structural and functional studies of these proteins and eventually lead to a better understanding of the whole machinery.

Of the vulnerability of orphan complex proteins: The case study of the E. coli IscU and IscS proteins

IANNUZZI, Clara;
2010

Abstract

IscS and IscU, the two central protein components of the iron sulfur cluster assembly machinery, form a complex that is still relatively poorly characterized. In an attempt to standardize the purification of these proteins for structural studies we have developed a protocol to produce them individually in high concentration and purity. We show that IscS is a rather robust protein as long as it is produced in a PLP loaded form and that this co-factor is essential for fold stability and enzyme activity. In contrast to previous evidence, we also propose that, in contrast with previous evidence, IscU is a thermodynamically stable protein with a well defined fold but, when produced in isolation, is a 'complex-orphan protein' that is prone to unfolding if not stabilised by a co-factor or a protein partner. Our work will facilitate further structural and functional studies of these proteins and eventually lead to a better understanding of the whole machinery.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11591/344851
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