| Summary: | 碩士 === 國立清華大學 === 生物資訊與結構生物研究所 === 98 === H+-translocating inorganic pyrophosphatase (H+-PPase), a
member of proton pumps, utilizes inorganic pyrophosphate (PPi) as energy source to generate proton-motive forces across membranes. H+-PPase from Clostridium tetani (CtH+-PPase) contains 16 transmembrane domains and shows only three tryptophan residues in entire protein. Tryptophan is an important aromatic amino acid for many proteins. Therefore, we investigated the role of tryptophan residues in CtH+-PPase by means of tryptophan-substitution mutagenesis. The PPi hydrolysis activities, proton translocations, and coupling efficiencies of the mutated CtH+-PPases were then determined. Substitution for W75 showed a dramatic decrease in coupling ratio, indicating obvious structural changes of the protein. Furthermore, the effect of common ions (K+, Ca2+, Na+, F-) added to these mutated CtH+-PPases were measured. Addition of Ca2+, Na+, F- and subtraction of K+ would diminish the PPi hydrolysis activities in wild type and mutants to a certain extent.
The function and conformational change of CtH+-PPase were
accordingly elucidated in this study.
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