PMID:11732924
| Citation |
Ghosh, S, Misra, A, Ozarowski, A, Stuart, C and Maki, AH (2001) Characterization of the tryptophan residues of Escherechia coli alkaline phosphatase by phosphorescence and optically detected magnetic resonance spectroscopy. Biochemistry 40:15024-30 |
|---|---|
| Abstract |
The phosphorescence and zero field optically detected magnetic resonance (ODMR) of the tryptophan (Trp) residues of alkaline phosphatase from Escherechia coli are examined. Each Trp is resolved optically and identified with the aid of the W220Y mutant and the terbium complex of the apoenzyme. Trp(109), known from earlier work to be the source of room-temperature phosphorescence (RTP), emits a highly resolved low-temperature phosphorescence (LTP) spectrum and has the narrowest ODMR bands observed thus far from any protein site, revealing a uniquely homogeneous local environment. The decay kinetics of Trp(109) at 1.2 K reveals that the major triplet population (70%) undergoes inefficient crystallike spin-lattice relaxation by direct interaction with lattice phonons, the remainder being relaxed efficiently by local disorder modes. The latter population is smaller than is typical for protein sites, suggesting an unusual degree of local rigidity and order consistent with the long-lived RTP. Trp(220) emits a broader LTP spectrum originating to the blue of Trp(109). It has typically broad ODMR bands consistent with local heterogeneity. The LTP of Trp(268) has an ill-defined origin blue shifted relative to Trp(220) and ODMR frequencies consistent with a greater degree of solvent exposure. Trp(268) has noticeable dispersion of its decay kinetics, consistent with quenching at the triplet level by a nearby disulfide residue. |
| Links | |
| Keywords |
Alkaline Phosphatase/chemistry; Alkaline Phosphatase/genetics; Bacterial Proteins/chemistry; Escherichia coli/enzymology; Escherichia coli/genetics; Luminescent Measurements; Magnetic Resonance Spectroscopy/methods; Molecular Structure; Temperature; Tryptophan/chemistry |
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