A quantitative assay to determine the amount of Signal Peptidase I in E. coli and the orientation of membrane vesicles

Abstract

The number of Signal Peptidase I (SPasel) molecules perE. coli cell was determined using western blot techniques. Different strains were found to contain approximately 1000 SPasel molecules per cell during exponential growth. Based on the activity of SPasel in vitro it could be estimated that this amount is sufficient to process all translocated precursors. SPasel did not appear to be under growth phase dependent control, but was constitutively expressed. The quantitative western blot technique was also used to establish the orientation and intactness of isolated inner membrane vesicles.

Key Words:

• Signal Peptidase I
• protein translocation
• quantitative western blotting
• membrane vesicle orientation