Role of the heme propionates in the interaction of heme with apomyoglobin and apocytochrome b(5)

被引:78
作者
Hunter, CL
Lloyd, E
Eltis, LD
Rafferty, SP
Lee, H
Smith, M
Mauk, AG
机构
[1] UNIV BRITISH COLUMBIA,PROT ENGN NETWORK CTR EXCELLENCE,VANCOUVER,BC V6T 1Z3,CANADA
[2] UNIV BRITISH COLUMBIA,BIOTECHNOL LAB,VANCOUVER,BC V6T 1Z3,CANADA
[3] UNIV GUELPH,DEPT ENVIRONM BIOL,GUELPH,ON N1G 2W1,CANADA
关键词
D O I
10.1021/bi961385u
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 heme propionate groups of both myoglobin (Mb) and cytochrome b(5) form hydrogen bonds with nearby surface amino acids residues that are believed to stabilize the heme-protein complex. To evaluate the magnitude of this stabilization, the kinetics of heme dissociation from variants of horse heart Mb and cytochrome b(5) in which these hydrogen bonding interactions have been systematically eliminated were studied by the method of Hargrove and colleagues (1994), and their thermal stability was assessed. Elimination of each hydrogen bond was found to decrease the thermal stability of the proteins and increase the rate constant for heme dissociation in a progressive fashion. For the Mb derivatives, H-1-NMR studies indicate that the elimination of individual hydrogen bonds also affects the rate at which the heme orientational equilibrium is achieved, In both types of kinetics experiment, the effects of decreasing the number of potential hydrogen bonding interactions are found to be cumulative, Despite their kinetic effects, elimination of these hydrogen bonding interactions had no influence on the initial distribution of heme orientational isomers immediately following reconstitution or on the equilibrium constant of heme orientational disorder. The interactions between the heme propionates and nearby protein residues play a partial role in the stabilization of the heme-protein complex and are a major factor in the kinetic ''trapping'' of the minor heme orientation. Comparisons of the various rate constants determined for the mechanism of heme binding and reorientation suggests that the intramolecular reorientation mechanism is slightly favored over the intermolecular mechanism.
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页码:1010 / 1017
页数:8
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