Backbone dynamics of a bacterially expressed peptide from the receptor binding domain of Pseudomonas aeruginosa pilin strain PAK from heteronuclear 1H-15N NMR spectroscopy

被引:19
作者
Campbell, AP [1 ]
Spyracopoulos, L
Irvin, RT
Sykes, BD
机构
[1] Univ Washington, Sch Pharm, Dept Med Chem, Seattle, WA 98195 USA
[2] Univ Alberta, Dept Biochem, Edmonton, AB T6G 2H7, Canada
[3] Univ Alberta, Dept Med Microbiol & Immunol, Edmonton, AB T6G 2H7, Canada
[4] Univ Alberta, Prot Engn Network Ctr Excellence, Edmonton, AB T6G 2S2, Canada
关键词
backbone dynamics; pilin; N-15 NMR relaxation; Pseudomonas aeruginosa; recombinant peptide;
D O I
10.1023/A:1008311319998
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 backbone dynamics of a N-15-labeled recombinant PAK pilin peptide spanning residues 128-144 in the C-terminal receptor binding domain of Pseudomonas aeruginosa pilin protein strain PAK (Lys(128)-Cys-Thr-Ser-Asp-Gln-Asp-Glu-Gln-Phe-Ile-Pro-Lys-Gly-Cys-Ser-Lys(144)) were probed by measurements of N-15 NMR relaxation. This PAK(128-144) sequence is a target for the design of a synthetic peptide vaccine effective against multiple strains of P. aeruginosa infection. The N-15 longitudinal (T-1) and transverse (T-2) relaxation rates and the steady-state heteronuclear {H-1}-N-15 NOE were measured at three fields (7.04, 11.74 and 14.1 Tesla), five temperatures (5, 10, 15, 20, and 25 degrees C ) and at pH 4.5 and 7.2. Relaxation data was analyzed using both the 'model-free' formalism [Lipari, G. and Szabo, A. (1982) J. Am. Chem. Soc., 104, 4546-4559 and 4559-4570] and the reduced spectral density mapping approach [Farrow, N.A., Szabo, A., Torchia, D.A. and Kay, L.E. (1995) J. Biomol. NMR, 6, 153-162]. The relaxation data, spectral densities and order parameters suggest that the type I and type II beta-turns spanning residues Asp(134)-Glu-Gln-Phe(137) and Pro(139)-Lys-Gly-Cys(142), respectively, are the most ordered and structured regions of the peptide. The biological implications of these results will be discussed in relation to the role that backbone motions play in PAK pilin peptide immunogenicity, and within the framework of developing a pilin peptide vaccine capable of conferring broad immunity across P. aeruginosa strains.
引用
收藏
页码:239 / 255
页数:17
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