Effects of protonation on the hydrolysis of triphosphate in vacuum and the implications for catalysis by nucleotide hydrolyzing enzymes. Issue 1 (December 2016)

• Record Type: Journal Article
• Authors: Kiani, Farooq; Fischer, Stefan
• Is Part Of: BMC biochemistry. Volume 17:Issue 1(2016)
• Page Start: 1
• Page End: 13
• Publication Date: 2016-12
• Subjects: ATP hydrolysis -- ATPase -- GTPase -- Enzymatic mechanism -- Concurrent and sequential mechanisms -- Computational biochemistry; Biochemistry -- Periodicals; 572.05
• DOI: 10.1186/s12858-016-0068-7
• Languages: English
• ISSNs: 1471-2091

Abstract

Abstract Background Nucleoside triphosphate (NTP) hydrolysis is a key reaction in biology. It involves breaking two very stable bonds (one P–O bond and one O–H bond of water), in either a concurrent or a sequential way. Here, we systematically examine how protonation of the triphosphate affects the mechanism of hydrolysis. Results The hydrolysis reaction of methyl triphosphate in vacuum is computed with protons in various numbers and position on the three phosphate groups. Protonation is seen to have a strong catalytic effect, with the reaction mechanism depending highly on the protonation pattern. Conclusion This dependence is apparently complicated, but is shown to obey a well-defined set of rules: Protonation of the α- and β-phosphate groups favors a sequential hydrolysis mechanism, whereas γ-protonation favors a concurrent mechanism, the two effects competing with each other in cases of simultaneous protonation. The rate-limiting step is always the breakup of the water molecule while it attacks the γ-phosphorus, and its barrier is lowered by γ-protonation. This step has significantly lower barriers in the sequential reactions, because the dissociated γ-metaphosphate intermediate (Pγ O3 − ) is a much better target for water attack than the un-dissociated γ-phosphate (−Pγ O4 2− ). The simple chemical logic behind these rules helps to better understand the catalytic strategy used by NTPase enzymes, as illustrated here for the catalytic pocket of myosin. A set of rules was determined that describes how protonating the phosphate groups affects the hydrolysis mechanism of methyl triphosphate: Protonation of the α- and/or β- phosphate groups promotes a sequential mechanism in which P-O bond breaking precedes the breakup of the attacking water, whereas protonation of the γ-phosphate promotes a concurrent mechanism and lowers the rate-limiting barrier of water breakup. The role played by individual protein residues in the catalytic pocket of triphosphate hydrolysing enzymes can be assigned accordingly. Graphical abstract