Activation mechanism of the CO sensor CooA - Mutational and resonance Raman spectroscopic studies

被引:43
作者
Coyle, CM
Puranik, M
Youn, H
Nielsen, SB
Williams, RD
Kerby, RL
Roberts, GP
Spiro, TG [1 ]
机构
[1] Princeton Univ, Dept Chem, Princeton, NJ 08544 USA
[2] Univ Wisconsin, Dept Bacteriol, Madison, WI 53706 USA
关键词
D O I
10.1074/jbc.M301000200
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
CooA is a CO-dependent heme protein transcription factor of the bacterium Rhodospirillum rubrum. CO binding to its heme causes CooA to bind DNA and activate expression of genes for CO metabolism. To understand the nature of CO activation, several CooA mutational variants have been studied by resonance Raman spectroscopy, in vivo activity measurements, and DNA binding assays. Analysis of the Fe-C and C-O stretching Raman spectroscopy bands permits the conclusion that when CO displaces the Pro(2) heme ligand, the protein forms a hydrophobic pocket in which the C-helix residues Gly(117), Leu(116), and Ile(113) are close to the bound CO. The displaced Pro(2) terminus is expelled from this pocket, unless the pH is raised above the pK(a), in which case the terminus remains in H- bond contact. The pK(a) for this transition is 8.6, two units below that of aqueous proline, reflecting the hydrophobic nature of the pocket. The proximal Fe - His bond in Fe[II] CooA is as strong as it is in myoglobin but is weakened by CO binding, an effect attributable to loss of an H- bond from the proximal His(77) ligand to the adjacent Asn(42) side chain. A structural model is proposed for the position of the CO-bound heme in the active form of CooA, which has implications for the mechanism of CO activation.
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收藏
页码:35384 / 35393
页数:10
相关论文
共 34 条
[1]  
Ausubel F. M., 1995, SHORT PROTOCOLS MOL, P21
[2]   TIME-RESOLVED RESONANCE RAMAN-SPECTROSCOPY OF CYTOCHROME-C REDUCED BY PULSE-RADIOLYSIS [J].
CARTLING, B ;
WILBRANDT, R .
BIOCHIMICA ET BIOPHYSICA ACTA, 1981, 637 (01) :61-68
[3]   CooA, CAP and allostery [J].
Chan, MK .
NATURE STRUCTURAL BIOLOGY, 2000, 7 (10) :822-824
[4]  
DESBOIS A, 1992, EUR BIOPHYS J BIOPHY, V20, P321, DOI 10.1007/BF00196591
[5]   SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modeling [J].
Guex, N ;
Peitsch, MC .
ELECTROPHORESIS, 1997, 18 (15) :2714-2723
[6]   Repositioning about the dimer interface of the transcription regulator CooA: A major signal transduction pathway between the effector and DNA-binding domains [J].
Kerby, RL ;
Youn, H ;
Thorsteinsson, MV ;
Roberts, GP .
JOURNAL OF MOLECULAR BIOLOGY, 2003, 325 (04) :809-823
[7]  
Kitagawa T., 1988, Biological Applications of Raman Spectroscopy, P97
[8]  
Lanzilotta WN, 2000, NAT STRUCT BIOL, V7, P876
[9]   IS BOUND CO LINEAR OR BENT IN HEME-PROTEINS - EVIDENCE FROM RESONANCE RAMAN AND INFRARED SPECTROSCOPIC DATA [J].
LI, XY ;
SPIRO, TG .
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 1988, 110 (18) :6024-6033
[10]   Fluorescence polarization analysis of protein-DNA and protein-protein interactions [J].
Lundblad, JR ;
Laurance, M ;
Goodman, RH .
MOLECULAR ENDOCRINOLOGY, 1996, 10 (06) :607-612