Biologjia Online

Characterization of an Integral Membrane Protein

Isoprenylcysteine carboxyl methyltransferase (Icmt) is an endoplasmic reticulum enzyme responsible for carboxylmethylation of prenylated proteins. This methylation is critical for the proper localization and functioning of Ras proteins, thus making it a potentially useful chemotherapeutic target for Ras-based cancers. However, Icmts have multiple membrane spanning domains, which present a challenge for their purification and characterization. In this paper, Jessica L. Anderson and colleagues purify an Icmt from S. cerevisiae (Ste14p) to homogeneity using high level expression of the His-tagged protein followed by solubilization in -D-dodecylmaltoside and metal affinity column chromatography. The purified enzyme retained activity and was able to methylate both a model substrate (N-acetyl-S-farnesyl-L-cysteine) and Ras. The authors also resolved the question as to whether the enzyme prefers farnesylated or geranylgeranylated substrates: it likes them both equally. These results pave the way for further characterization of this carboxyl methyltransferase as well as purification of other integral membrane proteins.

posted by Selfmaderadio @ 12:07 e pasdites