The pore size of the autotransporter domain is critical for the active translocation of the passenger domain.
The autotransporter mode of surface presentation in Gram-negative bacteria requires a hypothetical C-terminal beta-barrel which makes up an aqueous channel in the outer membrane. PalA is a Pseudomonas sp. autotransporter lipolytic protein. PalA is a 66 kDa protein that is composed of two parts, the N-terminal region (Ala(1)-Ala(296)) similar to the GDSL lipases and the C-terminal region (Leu(320)-Phe(612)) to the autotransporter. In this report, we provide biochemical and structural evidence demonstrating that the pore size of the beta-barrel conduit is important in delivering the N-terminal domain to the cell surface. Among all the autotransporter domains two strictly conserved residues (Pro(478) and Gly(576) in PalA) are converted to other various residues using site-directed mutagenesis. This investigation was made into the different pore-size mutants, affecting the folding of N-terminal domain. Wild beta-domain contains a cavity of approximately 2 nm diameter that is optimal for the active conformation of the N-terminal domains. However, deviation from the proper size of the pore, whether it is larger or smaller, is not suitable for the proper folding of the N-terminal catalytic domain.
Lee
Hyun Woo
HW
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1 Kusong-dong, Yuseong, Taejon 305-701, Republic of Korea.
Byun
Si Myung
SM
D000595:Amino Acid Sequence; D020134:Catalytic Domain; D010450:Endopeptidases / Q000378:metabolism; D008049:Lipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008066:Lipolysis; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018272:Porins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011549:Pseudomonas / Q000201:enzymology / Q000378:metabolism
D018272:Porins; D008049:Lipase; D010450:Endopeptidases