Literature summary for 3.6.1.1 extracted from

Oksanen, E.; Ahonen, A.K.; Tuominen, H.; Tuominen, V.; Lahti, R.; Goldman, A.; Heikinheimo, P.
A complete structural description of the catalytic cycle of yeast pyrophosphatase (2007), Biochemistry, 46, 1228-1239.

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant wild-type enzyme or mutants E48D, Y93F, D115E, D117E, D120E, D120N, and D152E in 60 mM MES, pH 6.0, and 10 mM Mg2+, 4°C, 0.008 ml sitting drops in the presence of 5 mM Mg2+, 1 mM PO43-, and a MPD concentration gradient from 16 to 19%, 2-4 weeks, cryoprotection by soaking of crystals at 4°C in 32% MPD, 30 mM MES, 10 mM Mg2+, and 1 mM PO43- for a few min, X-ray diffraction structure determination and analysis at 1.5-1.9 A resolution	Saccharomyces cerevisiae

Crystallization (Comment)

Organism

purified recombinant wild-type enzyme or mutants E48D, Y93F, D115E, D117E, D120E, D120N, and D152E in 60 mM MES, pH 6.0, and 10 mM Mg2+, 4°C, 0.008 ml sitting drops in the presence of 5 mM Mg2+, 1 mM PO43-, and a MPD concentration gradient from 16 to 19%, 2-4 weeks, cryoprotection by soaking of crystals at 4°C in 32% MPD, 30 mM MES, 10 mM Mg2+, and 1 mM PO43- for a few min, X-ray diffraction structure determination and analysis at 1.5-1.9 A resolution

Saccharomyces cerevisiae

Protein Variants

Protein Variants	Comment	Organism
D115E	site-directed mutagenesis, the mutation affects metal binding and the hydrogen bonding network in the active, in contrary to the wild-type enzyme, the mutant shows an open conformation variant of the hitherto unobserved two-phosphate and two bridging water active site, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae
D117E	site-directed mutagenesis, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae
D120E	site-directed mutagenesis, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae
D120N	site-directed mutagenesis, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae
D152E	site-directed mutagenesis, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae
E48D	site-directed mutagenesis, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae
Y93F	site-directed mutagenesis, the mutation affects metal binding and the hydrogen bonding network in the active, crystal structure determination with bound phosphate and Mg2+, and comparison to the wild-type enzyme structure	Saccharomyces cerevisiae

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	metal binding affinities and kinetics of wild-type and mutant enzymes, overview	Saccharomyces cerevisiae

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	the pyrophosphatase requires four divalent metal cations for catalysis, magnesium provides the highest activity	Saccharomyces cerevisiae

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
diphosphate + H2O	Saccharomyces cerevisiae	-	2 phosphate	-	r

Organism

Organism	UniProt	Comment	Textmining
Saccharomyces cerevisiae	P00817	-	-

Reaction

Reaction	Comment	Organism	Reaction ID
diphosphate + H2O = 2 phosphate	substrate binding structure and catalytic mechanism	Saccharomyces cerevisiae	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
diphosphate + H2O	-	Saccharomyces cerevisiae	2 phosphate	-	r
diphosphate + H2O	the rate-determining step for the forward reaction with Mg2+ is hydrolysis of PPi, the wild-type active site shows a closed comformation with one of the two product phosphates already dissociated, active site residues Tyr93 and Asp115 are important, six-state catalytic mechanism, overview	Saccharomyces cerevisiae	2 phosphate	in the reverse, net synthesis direction, the rate-determining step is not the condensation of the two phosphate ions but the previous step, which involves isomerization of the enzyme	r

Synonyms

Synonyms	Comment	Organism
inorganic pyrophosphatase	-	Saccharomyces cerevisiae
PPase	-	Saccharomyces cerevisiae