Human CaaX protease ZMPSTE24 expressed in yeast: Structure and inhibition by HIV protease inhibitors.

The function and localization of proteins and peptides containing C-terminal "CaaX" (Cys-aliphatic-aliphatic-anything) sequence motifs are modulated by post-translational attachment of isoprenyl groups to the cysteine sulfhydryl, followed by proteolytic cleavage of the aaX amino acids. The zinc metalloprotease ZMPSTE24 is one of two enzymes known to catalyze this cleavage. The only identified target of mammalian ZMPSTE24 is prelamin A, the precursor to the nuclear scaffold protein lamin A.

Structure of the integral membrane protein CAAX protease Ste24p.

Posttranslational lipidation provides critical modulation of the functions of some proteins. Isoprenoids (i.e.

Purification of transmembrane proteins from Saccharomyces cerevisiae for X-ray crystallography.

To enhance the quantity and quality of eukaryotic transmembrane proteins (TMPs) available for structure determination by X-ray crystallography, we have optimized protocols for purification of TMPs expressed in the yeast Saccharomyces cerevisiae. We focused on a set of the highest-expressing endogenous yeast TMPs for which there are established biochemical assays. Genes encoding the target TMPs are transferred via ligation-independent cloning to a series of vectors that allow expression of reading frames fused to C-terminal His10 and ZZ (IgG-binding) domains that are separated from the reading frame by a cleavage site for rhinovirus 3C protease.