期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 106, 期 45, 页码 18984-18989出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0905433106
关键词
laser flash photolysis; reaction kinetics; silica gel
资金
- MiUR
- Fondazione Cariparma fellowship
- Fund for Scientific Research (FWO)
Neuroglobin (Ngb), a member of the globin superfamily, was found in the brain of vertebrates and is suggested to play a neuroprotective function under hypoxic conditions by scavenging nitrogen monoxide ( NO) through a dioxygenase activity. In order for such a reaction to efficiently take place and to minimize the release of reactive intermediates in the cytosol, the cosubstrates O-2 and NO and other unstable reaction intermediates should bind sequentially to docking sites in the protein matrix. We have characterized the accessibility of these sites by analyzing the geminate CO rebinding kinetics to the heme moiety observed upon nanosecond flash photolysis of the Ngb-CO complex encapsulated in silica gels. The geminate rebinding phase showed a remarkable complexity, revealing the presence of a system of secondary docking sites where ligands are stored for hundreds of microseconds. Most kinetics steps display little temperature dependence, demonstrating that ligands can easily migrate through the cavities, except for the slowest reaction intermediate, possibly reflecting a structural conformational change reshaping the system of cavities. This conformational change is unrelated with distal His E7 binding to the heme, as it persists for the HE7L mutant. Overall, data are consistent with the presence of a discrete system of docking sites, possibly acting as reservoirs for the putative cosubstrates and for other reactive species involved in the physiologically relevant reaction.
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