title: The use of sarkosyl in generating soluble protein after bacterial expression.
abstract: Actin, like many other proteins, is highly insoluble after expression in Escherichia coli. In order to understand the origin of insoluble aggregates, we asked whether morphological inclusions were always correlated with insolubility. The strain expressing actin was compared to one that expresses part of the myosin tail; the latter strain yields soluble protein after various cell lysis or disruption procedures. Morphological inclusions were observed in both strains, indicating there is no obligate relationship between solubility and inclusions. Studies presented here suggest that extreme insolubility results from coaggregation of the actin with bacterial outer membrane components upon bacterial lysis. The properties of the outer membrane have been exploited in the development of nondenaturing procedures that yield soluble actin. One procedure involves the disruption of coaggregates with sarkosyl detergent (N-laurylsarcosine); another prevents the formation of coaggregates by lysing in the presence of sarkosyl. These methods may be useful for other proteins that become insoluble after bacterial expression.
authors.affiliation: Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, NY 10461.
authors.forename: S
authors.initials: S
authors.lastname: Frankel
authors.forename: R
authors.initials: R
authors.lastname: Sohn
authors.forename: L
authors.initials: L
authors.lastname: Leinwand
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000302:isolation & purification; D015153:Blotting, Western; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000648:ultrastructure; D008854:Microscopy, Electron; D008970:Molecular Weight; D009218:Myosins / Q000235:genetics / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000302:isolation & purification*; D012521:Sarcosine / Q000031:analogs & derivatives; D012995:Solubility
chemical_list: D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D011994:Recombinant Proteins; C025231:sarkosyl; D009218:Myosins; D012521:Sarcosine