Crystals of Tryptophan Indole-Lyase and Tyrosine Phenol-Lyase Form Stable Quinonoid Complexes
| Autor: | Stefano Bruno, Luca Ronda, Andrea Mozzarelli, Tatyana V. Demidkina, Robert S. Phillips, Lyudmila N. Zakomirdina |
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| Rok vydání: | 2002 |
| Předmět: |
Conformational change
Aldimine Ultraviolet Rays Stereochemistry Tyrosine phenol-lyase Photochemistry Biochemistry Catalysis Enzyme catalysis Serine Proteus vulgaris Tyrosine Tyrosine Phenol-Lyase Molecular Biology chemistry.chemical_classification biology Tryptophanase Tryptophan Cell Biology biology.organism_classification Citrobacter freundii Dissociation constant Kinetics Models Chemical chemistry Spectrophotometry Crystallization Protein Binding |
| Zdroj: | Journal of Biological Chemistry. 277:21592-21597 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m200216200 |
| Popis: | The binding of substrates and inhibitors to wild-type Proteus vulgaris tryptophan indole-lyase and to wild type and Y71F Citrobacter freundii tyrosine phenol-lyase was investigated in the crystalline state by polarized absorption microspectrophotometry. Oxindolyl-lalanine binds to tryptophan indole-lyase crystals to accumulate predominantly a stable quinonoid intermediate absorbing at 502 nm with a dissociation constant of 35 microm, approximately 10-fold higher than that in solution. l-Trp or l-Ser react with tryptophan indole-lyase crystals to give, as in solution, a mixture of external aldimine and quinonoid intermediates and gem-diamine and external aldimine intermediates, respectively. Different from previous solution studies (Phillips, R. S., Sundararju, B.,Faleev, N. G. (2000) J. Am. Chem. Soc. 122, 1008-1114), the reaction of benzimidazole and l-Trp or l-Ser with tryptophan indole-lyase crystals does not result in the formation of an alpha-aminoacrylate intermediate, suggesting that the crystal lattice might prevent a ligand-induced conformational change associated with this catalytic step. Wild-type tyrosine phenol-lyase crystals bind l-Met and l-Phe to form mixtures of external aldimine and quinonoid intermediates as in solution. A stable quinonoid intermediate with lambda(max) at 502 nm is accumulated in the reaction of crystals of Y71F tyrosine phenol-lyase, an inactive mutant, with 3-F-l-Tyr with a dissociation constant of 1 mm, approximately 10-fold higher than that in solution. The stability exhibited by the quinonoid intermediates formed both by wild-type tryptophan indole-lyase and by wild type and Y71F tyrosine phenol-lyase crystals demonstrates that they are suitable for structural determination by x-ray crystallography, thus allowing the elucidation of a key species of pyridoxal 5'-phosphate-dependent enzyme catalysis. |
| Databáze: | OpenAIRE |
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