Porin isolated from the outer membrane of Erwinia amylovora and its encoding gene.
A major Erwinia amylovora outer-membrane protein (Omp-EA) and the gene encoding for this protein (omp-EA) were isolated and characterized. The native Omp-EA protein forms a trimeric structure of approximately 114 kDa. This protein demonstrated high resistance to detergents such as SDS and octyl-glucopyranoside, but disaggregated to monomers with a molecular weight (MW) of approximately 39 kDa after heating at 95 degrees C for 10 minutes in sample buffer. The pore-forming ability of the oligomeric Omp-EA was determined by the liposome swelling assay, demonstrating that the oligomeric protein formed nonspecific channels with an exclusion limit of approximately 660 Da. On dissociation, the monomers did not exhibit pore-forming ability. The omp-EA gene was cloned and sequenced (GenBank Accession No. DQ184680). Sequence analysis revealed an open reading frame of 1152 bases. The deduced amino-acid sequence had 383 amino acids. The mature protein consisted of 362 amino acids and had a calculated MW of 39,210 Da. Multiple-sequence alignment of Omp-EA with other porins from the Enterobacteriaceae family revealed 51% to 63% identity. The first 16 amino acids from the N-terminal exhibited the highest identity (100%) to the porins OmpC, OmpF, and PhoE of Escherichia coli. Two methods were used to predict the secondary structure: APSSP2 and Hidden and Markov's model. The monomers of Omp-EA porin presented a topology of 16 transmembranal beta-strands. The area of the loops between the beta -strands was proposed. It is suggested that further research on the porin and its loops may be important for understanding the mechanism of E. amylovor to invade plant tissues.
Elazar
M
M
The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, 52900, Israel.
Halfon
D
D
Pechatnikov
I
I
Nitzan
Y
Y
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D044084:Erwinia amylovora / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D018272:Porins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017433:Protein Structure, Secondary; D017422:Sequence Analysis, DNA
D001425:Bacterial Outer Membrane Proteins; D018272:Porins