Literature summary for 2.8.2.20 extracted from

Tanaka, S.; Nishiyori, T.; Kojo, H.; Otsubo, R.; Tsuruta, M.; Kurogi, K.; Liu, M.C.; Suiko, M.; Sakakibara, Y.; Kakuta, Y.
Structural basis for the broad substrate specificity of the human tyrosylprotein sulfotransferase-1 (2017), Sci. Rep., 7, 8776 .

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of N-terminally His6-tagged TPST1 (Lys43-Glu370) in Escherichia coli strain Origami (DE3) carrying the pGro7 plasmid encoding chaperonin proteins	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified TPST1 complexed with two substrate peptides, C4 complement-deived C4P5Y5 or a gastrin peptide, and PAP. PST1-PAP-C4P5Y5 is crystallized by sitting drop vapor diffusion method, mixing of 2.5 mg/ml protein, 2 mM PAP, and 1 mM C4P5Y5 peptide in 50 mM Tris-HCl, pH 7.0, and 200 mM NaCl with reservoir solution consisting of 0.2 M trimethylamine N-oxide dihydrate, 0.1 M Tris-HCl, pH 8.5, and 20% w/v PEG monomethyl ether 2000, nine months, at 20°C. TPST1-PAP-gastrin peptide is crystallized by sitting drop vapor diffusion method, mixing of 5.0 mg/ml protein, 2 mM PAP, and 3 mM gastrin peptide in 50 mM Tris-HCl, pH 7.0, and 200 mM NaCl with reservoir solution consisting of 0.2 M potassium sodium tartrate trihydrate and 19.5% w/v PEG 3350, two months, at 20°C. X-ray diffraction structure determination and analysis at 1.6 A and 2.3 A resolution, respectively. The asymmetric unit of human TPST1-PAP-C4P5Y5 contains two human TPST1 molecules, which assemble to form a dimer. In contrast, the asymmetric unit of TPST1-PAP-gastrin peptide contains four human TPST1 molecules, which assemble to form two dimers	Homo sapiens

Crystallization (Comment)

Organism

purified TPST1 complexed with two substrate peptides, C4 complement-deived C4P5Y5 or a gastrin peptide, and PAP. PST1-PAP-C4P5Y5 is crystallized by sitting drop vapor diffusion method, mixing of 2.5 mg/ml protein, 2 mM PAP, and 1 mM C4P5Y5 peptide in 50 mM Tris-HCl, pH 7.0, and 200 mM NaCl with reservoir solution consisting of 0.2 M trimethylamine N-oxide dihydrate, 0.1 M Tris-HCl, pH 8.5, and 20% w/v PEG monomethyl ether 2000, nine months, at 20°C. TPST1-PAP-gastrin peptide is crystallized by sitting drop vapor diffusion method, mixing of 5.0 mg/ml protein, 2 mM PAP, and 3 mM gastrin peptide in 50 mM Tris-HCl, pH 7.0, and 200 mM NaCl with reservoir solution consisting of 0.2 M potassium sodium tartrate trihydrate and 19.5% w/v PEG 3350, two months, at 20°C. X-ray diffraction structure determination and analysis at 1.6 A and 2.3 A resolution, respectively. The asymmetric unit of human TPST1-PAP-C4P5Y5 contains two human TPST1 molecules, which assemble to form a dimer. In contrast, the asymmetric unit of TPST1-PAP-gastrin peptide contains four human TPST1 molecules, which assemble to form two dimers

Homo sapiens

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
0.00767	-	C4P5Y5 peptide	recombinant enzyme TPST1, pH 6.0, 30°C	Homo sapiens
0.652	-	gastrin peptide	recombinant enzyme TPST1, pH 6.0, 30°C	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
3'-phosphoadenylyl sulfate + protein tyrosine	Homo sapiens	-	adenosine 3',5'-bisphosphate + protein tyrosine-O-sulfate	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	O60507	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant N-terminally His6-tagged TPST1 (Lys43-Glu370) from Escherichia coli strain Origami (DE3) by nickel affinity chromatography, ultrafiltration and gel filtration, the tag is cleaved off through thrombin, thrombin is removed by benzamidine affinity chromatography, followed by desalting gel filtration	Homo sapiens

Reaction

Reaction	Comment	Organism	Reaction ID
3'-phosphoadenylyl sulfate + protein tyrosine = adenosine 3',5'-bisphosphate + protein tyrosine-O-sulfate	catalytic mechanism, structure-function analysis	Homo sapiens	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
3'-phosphoadenylyl sulfate + C4P5Y5 peptide	a C4 complement derived peptide substrate, sulfation of the Tyr741 residue	Homo sapiens	adenosine 3',5'-bisphosphate + sulfated C4P5Y5 peptide	-	?
3'-phosphoadenylyl sulfate + gastrin peptide	-	Homo sapiens	adenosine 3',5'-bisphosphate + sulfated gastrin peptide	-	?
3'-phosphoadenylyl sulfate + protein tyrosine	-	Homo sapiens	adenosine 3',5'-bisphosphate + protein tyrosine-O-sulfate	-	?
additional information	the structures of TPST1 complexed with two substrate peptides (C4P5Y5 derived from complement C4 and a gastrin peptide), that are catalysed by human TPST1 with significantly different efficiencies, reveal details about the binding modes found in the two complexes and into the sulfation mechanism for these substrates. The structures also contain only lumenal portion of the enzyme, without the N-terminus and the TM domain. The active site of human TPST1 recognizes the residue at the -3 position in the substrate through electrostatic and hydrophobic interactions. The enzyme shows broad substrate specificity	Homo sapiens	?	-	-

Subunits

Subunits	Comment	Organism
homodimer	human TPST1 forms a homodimer mediated by three consecutive alpha-helices (alpha2, alpha3 and alpha4)	Homo sapiens
More	the TPST1 catalytic domain comprises a single alpha/beta motif with a five-stranded parallel beta-sheet, flanked on both sides by alpha helices, and this structure is consistent with human TPST2 structure	Homo sapiens

Synonyms

Synonyms	Comment	Organism
TPST	-	Homo sapiens
TPST1	-	Homo sapiens
tyrosylprotein sulfotransferase	-	Homo sapiens
tyrosylprotein sulfotransferase-1	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Homo sapiens

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
0.00011	-	gastrin peptide	recombinant enzyme TPST1, pH 6.0, 30°C	Homo sapiens
0.00082	-	C4P5Y5 peptide	recombinant enzyme TPST1, pH 6.0, 30°C	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
6	-	assay at	Homo sapiens

General Information

General Information	Comment	Organism
evolution	in humans, there are only two TPST isoforms, designated TPST1 and TPST2	Homo sapiens
additional information	the TPST1 catalytic domain comprises a single alpha/beta motif with a five-stranded parallel beta-sheet, flanked on both sides by alpha helices, and this structure is consistent with human TPST2 structure. Structure-function analysis, overview. The 5'-phosphosulfate-binding (5'-PSB) motif, which is contained within a strand-loop-helix consisting of beta3 and alpha1, is central to this structural motif. Moreover, beta6 and alpha7 are also key elements that include the 3'-phosphate-binding (3-PB) motif. The positions of the catalytic residues R79, E100, K159 and S286 in human TPST1 are almost identical to those of the catalytic residues R78, E99, K158 and S285 in human TPST2, suggesting that the catalytic mechanism of human TPST1 is basically same to that of human TPST2	Homo sapiens
physiological function	tyrosylprotein sulfotransferases (TPSTs) are enzymes that catalyze post-translational tyrosine sulfation of proteins	Homo sapiens

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
0.00017	-	gastrin peptide	recombinant enzyme TPST1, pH 6.0, 30°C	Homo sapiens
0.107	-	C4P5Y5 peptide	recombinant enzyme TPST1, pH 6.0, 30°C	Homo sapiens