Characterization of a Half-Apo Derivative of Peptidylglycine Monooxygenase. Insight into the Reactivity of Each Active Site Copper
| Autor: | Ninian J. Blackburn, Shulamit Jaron |
|---|---|
| Rok vydání: | 2001 |
| Předmět: |
Spectrophotometry
Infrared Stereochemistry Peptidylglycine monooxygenase chemistry.chemical_element Peptide binding CHO Cells Biochemistry Mixed Function Oxygenases Substrate Specificity chemistry.chemical_compound Apoenzymes Oxygen Consumption Multienzyme Complexes Cricetinae Animals Reactivity (chemistry) Binding site Carbon Monoxide Binding Sites biology Spectrum Analysis X-Rays Active site Substrate (chemistry) Copper chemistry biology.protein Oligopeptides Derivative (chemistry) |
| Zdroj: | Biochemistry. 40:6867-6875 |
| ISSN: | 1520-4995 0006-2960 |
| Popis: | A derivative of peptidylglycine monooxygenase which lacks the CuH center has been prepared and characterized. This form of the enzyme is termed the half-apo protein. Copper-to-protein stoichiometric measurements establish that the protein binds only one of the two copper centers (CuM and CuH) found in the native enzyme. Confirmation that the methionine-containing CuM has been retained has been obtained from EXAFS experiments which show that the characteristic signature of the Cu-S(Met) interaction is preserved. The half-apo derivative binds 1 equiv of CO per copper with an IR frequency of 2092 cm(-1), and this monocarbonyl also displays the Cu-S(Met) interaction in its EXAFS spectrum. These results allow unambiguous assignment of the 2092 cm(-1) band as a CuM-CO species. Binding of CO in the presence of peptide substrate was also investigated. In the native enzyme, substrate induced binding of a second CO molecule with an IR frequency of 2062 cm(-1), tentatively assigned to a CO complex of the histidine-containing CuH site. Unexpectedly, this reactivity is also observed in the half-apo derivative, although the intensity distribution of the CO stretches now indicates that the copper has been partially transferred to a second site, believed to be CuH. The implications of this observation are discussed in terms of a possible additional peptide binding site close to the CuH center. |
| Databáze: | OpenAIRE |
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