Study of the ability of Agrobacterial protein VirE2 to form pores in membranes

Abstract

Experimental and computer-assisted studies of the ability of the Agrobacterium virulence protein VirE2 to interact with an artificial bilayer lipid membrane were carried out. The lipid mixture of 63.5% diphytanoyl phosphatidylcholine, 30% diphytanoyl phosphatidylethanolamine, and 6.5% diphytanoyl phosphatidylglycerol proved to be optimal for preparation of membranes that were stable for 20 min. When a field of 10 to 50 mV was applied, the conductance of the planar bilayer lipid membranes upon introduction of the recombinant protein VirE2 abruptly increased, indicating possible formation of single long-living (1.5–5 s) pores. No proteins homologous to the protein VirE2 from Agrobacterium tumefaciens (no. P08062) were found in the SWISS-PROT or NCBI databases. Fifteen β-sheets and 12α-helices were predicted for the protein VirE2 using PROFsec. Computer-aided methods were used to build model structures consisting of two and four VirE2 proteins. It has been shown for the first time that pores with the channel diameters of 2.2 or 4 nm can be formed in a model structure consisting of two or four VirE2 molecules, respectively, which is located in the bilayer membrane. The ends of a motile interdomain loop exposed in the channel formed by two proteins narrow the channel bore to 0.7 nm.