1998 Fiscal Year Final Research Report Summary
A Novel Interaction between a Host and a Guest in the Enzyme Active Site
Project/Area Number |
09680578
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Research Category |
Grant-in-Aid for Scientific Research (C)
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Allocation Type | Single-year Grants |
Section | 一般 |
Research Field |
Bioorganic chemistry
|
Research Institution | Kanagawa University |
Principal Investigator |
SAKURAI Tadamitsu Kanagawa Univ., Department of Applied Chem., Professor, 工学部, 教授 (10133456)
|
Project Period (FY) |
1997 – 1998
|
Keywords | Enzyme Active Site / papaiu / alpha-chymotrypsin / Chemical modification / Aromatie acyl pendant / Host-guest Interaction |
Research Abstract |
1-(Bromoacetyl)pyrene (BAP) was found to selectively alkylate the cysteine-25 residue (Cys-25) of papain, inactivating the enzyme. A comparison of the spectroscopic behavior of BAP-modified papain with that of the corresponding model compound showed that the pyrenecarbonylmethyl pendant (covalently bound to Cys-25 in a 1 : 1 stoichiometric ratio) gives a new absorption around 390 nm and a long-wavelength fluorescence at 465 nm, which are both characteristic of an associated dimer. On the other hand, spectroscopic analysis of the reactions of papain with bromoacetyl-substituted naphthalenes, which alkylate Cys-25 residue of this enzyme, allows us to propose that the charge-transfer complex formed between the naphthoylmethyl pendant and the histidine-159 residue is responsible for the appearance of a new long-wavelength absorption band. Hammett and ^<13>C NMR analyses of proton-dissociation processes for sulfonium-salt pendants on alpha chymotrypsin (ChT) which had been modified by 2-brom
… More
oacetophenone and its derivatives (as well as for their corresponding model sulfonium salts) show that a hydrophobic environment around the pendants (attached to the methionine-192 residue) destabilizes the sulfonium-salt form so as to substantially promote its proton-dissociation reaction. This produces a sulfonium ylide with a keto structure. The fluorogenic bromoacetyl-substituted coumarin derivatives 1 were also found to selectively alkylate the methionine-192 residue of ChT in a 1 : 1 stoichiometric ratio, inactivating this enzyme to a different extent. A comparison of pH-dependent fluorescence spectra of 1-modified ChT with those of the corresponding model sulfonium salt confirms that either a hydrophobic or ahydrophilic environment around the sulfonium-salt reporter groups on the derivatized enzyme affects the pKa of these groups in the excited singlet state to, if any, only a very small extent. It was proposed that the immobilized water molecules that are present in a hydrophilic environment of the ChT active site play a key role in causing the negligible change in pKa of the 1-derived pendants. Less
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Research Products
(6 results)