Literature summary for 1.14.14.9 extracted from

Kim, S.H.; Hisano, T.; Takeda, K.; Iwasaki, W.; Ebihara, A.; Miki, K.
Crystal structure of the oxygenase component (HpaB) of the 4-hydroxyphenylacetate 3-monooxygenase from Thermus thermophilus HB8 (2007), J. Biol. Chem., 282, 33107-33117.

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Crystallization (Commentary)

Crystallization (Comment)	Organism
crystal structures of ligand-free form, a binary complex with FAD, and a ternary complex with FAD and 4-hydroxyphenylacetate, to 2.0, 1.66 and 1.66 A resolution, respectively. Binding and dissociation of flavin are accompanied by conformational changes of the loop between beta5 and beta6 and of the loop between beta8 and beta9, leading to preformation of part of the substrate-binding site Ser197 and Thr198. The latter loop further changes its conformation upon binding of 4-hydroxyphenylacetate and obstructs the active site from the bulk solvent. Arg100 is located adjacent to the putative oxygen-binding site	Thermus thermophilus
HpaB in three states: a ligand-free form, a binary complex with FAD, and a ternary complex with FAD and 4-hydroxyphenylacetate, mixing of 0.004 ml of protein solution containing 5 mg/ml protein and 1 mM DTT, with an equal volume of reservoir solution containing 1.5 M ammonium sulfate, 0.1 M Tris-HCl, pH 8.5, and 25% v/v glycerol, crystals appear within a few min and grow during 1-4 days to final size, complex crystal formation by soaking of crystals in 5 mM ligand containing solutions, X-ray diffraction structure determination and analysis at 1.66-2.07 A resolution	Thermus thermophilus

Crystallization (Comment)

Organism

crystal structures of ligand-free form, a binary complex with FAD, and a ternary complex with FAD and 4-hydroxyphenylacetate, to 2.0, 1.66 and 1.66 A resolution, respectively. Binding and dissociation of flavin are accompanied by conformational changes of the loop between beta5 and beta6 and of the loop between beta8 and beta9, leading to preformation of part of the substrate-binding site Ser197 and Thr198. The latter loop further changes its conformation upon binding of 4-hydroxyphenylacetate and obstructs the active site from the bulk solvent. Arg100 is located adjacent to the putative oxygen-binding site

Thermus thermophilus

HpaB in three states: a ligand-free form, a binary complex with FAD, and a ternary complex with FAD and 4-hydroxyphenylacetate, mixing of 0.004 ml of protein solution containing 5 mg/ml protein and 1 mM DTT, with an equal volume of reservoir solution containing 1.5 M ammonium sulfate, 0.1 M Tris-HCl, pH 8.5, and 25% v/v glycerol, crystals appear within a few min and grow during 1-4 days to final size, complex crystal formation by soaking of crystals in 5 mM ligand containing solutions, X-ray diffraction structure determination and analysis at 1.66-2.07 A resolution

Thermus thermophilus

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
4-hydroxyphenylacetate + NAD(P)H + H+ + O2	Thermus thermophilus	-	3,4-dihydroxyphenylacetate + NAD(P)+ + H2O	-	?
4-hydroxyphenylacetate + NAD(P)H + H+ + O2	Thermus thermophilus HB8 / ATCC 27634 / DSM 579	-	3,4-dihydroxyphenylacetate + NAD(P)+ + H2O	-	?

Organism

Organism	UniProt	Comment	Textmining
Thermus thermophilus	-	-	-
Thermus thermophilus	Q5SJP8	-	-
Thermus thermophilus HB8 / ATCC 27634 / DSM 579	-	-	-
Thermus thermophilus HB8 / ATCC 27634 / DSM 579	Q5SJP8	-	-

Reaction

Reaction	Comment	Organism	Reaction ID
4-hydroxyphenylacetate + FADH2 + O2 = 3,4-dihydroxyphenylacetate + FAD + H2O	structural basis of the catalytic mechanism of HpaB	Thermus thermophilus	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
4-hydroxyphenylacetate + NAD(P)H + H+ + O2	-	Thermus thermophilus	3,4-dihydroxyphenylacetate + NAD(P)+ + H2O	-	?
4-hydroxyphenylacetate + NAD(P)H + H+ + O2	4-hydroxyphenylacetate, FAD, NADH, and O2 binding structures, overview, the binding and dissociation of flavin are accompanied by conformational changes of the loop between beta5 and beta6 and of the loop between beta8 and beta9, leading to preformation of part of the substrate-binding site involving Ser197 and Thr198. The latter loop further changes its conformation upon binding of 4-hydroxyphenylacetate and obstructs the active site from the bulk solvent. Arg100 is located adjacent to the putative oxygen-binding site and may be involved in the formation and stabilization of the C4a-hydroperoxyflavin intermediate	Thermus thermophilus	3,4-dihydroxyphenylacetate + NAD(P)+ + H2O	-	?
4-hydroxyphenylacetate + NAD(P)H + H+ + O2	-	Thermus thermophilus HB8 / ATCC 27634 / DSM 579	3,4-dihydroxyphenylacetate + NAD(P)+ + H2O	-	?
4-hydroxyphenylacetate + NAD(P)H + H+ + O2	4-hydroxyphenylacetate, FAD, NADH, and O2 binding structures, overview, the binding and dissociation of flavin are accompanied by conformational changes of the loop between beta5 and beta6 and of the loop between beta8 and beta9, leading to preformation of part of the substrate-binding site involving Ser197 and Thr198. The latter loop further changes its conformation upon binding of 4-hydroxyphenylacetate and obstructs the active site from the bulk solvent. Arg100 is located adjacent to the putative oxygen-binding site and may be involved in the formation and stabilization of the C4a-hydroperoxyflavin intermediate	Thermus thermophilus HB8 / ATCC 27634 / DSM 579	3,4-dihydroxyphenylacetate + NAD(P)+ + H2O	-	?

Subunits

Subunits	Comment	Organism
More	the enzyme consists of two components, an oxygenase HpaB and a reductase HpaC, structure analysis, overview	Thermus thermophilus

Cofactor

Cofactor	Comment	Organism	Structure
FAD	binding structure, binding causes conformational changes, overview	Thermus thermophilus
NAD(P)H	-	Thermus thermophilus