Interaction of a bacterially expressed peptide from the receptor binding domain of Pseudomonas aeruginosa pili strain PAK with a cross-reactive antibody:: Conformation of the bound peptide

被引:14
作者
Campbell, AP [1 ]
Wong, WY
Irvin, RT
Sykes, BD
机构
[1] Univ Washington, Sch Pharm, Dept Med Chem, Seattle, WA 98195 USA
[2] Univ Alberta, Prot Engn Network Ctr Excellence, Edmonton, AB T6G 2S2, Canada
[3] Univ Alberta, Dept Med Microbiol & Immunol, Edmonton, AB T6G 2H7, Canada
[4] Univ Alberta, Dept Biochem, Edmonton, AB T6G 2H7, Canada
关键词
D O I
10.1021/bi0016568
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The C-terminal receptor binding region of Pseudomonas aeruginosa pilin protein strain PAK (residues 128-144) has been the target for the design of a vaccine effective against P. aeruginosa infections. We have recently cloned and expressed a N-15-labeled PAK pilin peptide spanning residues 128-144 of the PAK pilin protein. The peptide exists as a major (trans) and minor (cis) species in solution, arising from isomerization around a central Ile(138)-Pro(139) peptide bond. The trans isomer adopts two well-defined turns in solution, a type I beta -turn spanning Asp(134)-Glu-Gln-Phe(137) and a type II beta -turn spanning Pro(139)-Lys-Gly-Cys(142). The cia isomer adopts only one well-defined type II beta -turn spanning Pro(139)-Lys-Gly-Cys(142) but displays evidence of a less ordered turn spanning Asp(132)-Gln-Asp-Glu(135). These turns have been implicated in cross-reactive antibody recognition. N-15-edited NMR spectroscopy was used to study the binding of the N-15-labeled PAK pilin peptide to an Fab fragment of a cross-reactive monoclonal antibody, PAK-13, raised against the intact PAK pilus. The results of these studies are as follows: the trans and cis isomers bind with similar affinity to the Fab, despite their different topologies; both isomers maintain the conformational integrity of their beta -turns when bound; binding leads to the preferential stabilization of the first turn over the second turn in each isomer; and binding leads to the perturbation of resonances within regions of the trans and cia backbone that undergo microsecond to millisecond motions. These slow motions may play a role in induced fit binding of the first turn to Fab PAK-13, which would allow the same antibody combining site to accommodate either trans or cis topology. More importantly for vaccine design, these motions may also play a role in the development of a broad-spectrum vaccine capable of generating an antibody therapeutic effective against the multiple strains of P. aeruginosa.
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页码:14847 / 14864
页数:18
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