PDBsum entry: 2ofw

Transferase

PDB id: 2ofw

Contents

Protein chains

(+ 2 more) 200 a.a. *

Ligands

ADX ×16

Metals

_MG ×6

Waters ×832

* Residue conservation analysis

PDB id:

2ofw

Name:

Transferase

Title:

Crystal structure of the apsk domain of human papss1 complexed with 2 aps molecules

Structure:

Aps kinase domain of the paps synthetase 1. Chain: a, b, c, d, e, f, g, h. Fragment: aps kinse domain (residues 1-227). Engineered: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: papss1, atpsk1, papss. Expressed in: escherichia coli. Expression_system_taxid: 562. Cutting site

Resolution:

2.05Å R-factor: 0.223 R-free: 0.282

Authors:

N Sekulic,A Lavie

Key ref:

N.Sekulic et al. (2007). Elucidation of the active conformation of the APS-kinase domain of human PAPS synthetase 1. J Mol Biol, 367, 488-500. PubMed id: 17276460 DOI: 10.1016/j.jmb.2007.01.025

Date:

04-Jan-07 Release date: 10-Apr-07

Protein chains

O43252  (PAPS1_HUMAN) -  Bifunctional 3'-phosphoadenosine 5'-phosphosulfate synthase 1

Seq: 624 a.a.

Struc: 200 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class 2: E.C.2.7.1.25 - Adenylyl-sulfate kinase.
• Reaction: ATP + adenylyl sulfate = ADP + 3'-phosphoadenylyl sulfate

ATP + adenylyl sulfate (Bound ligand (Het Group name = ADX) corresponds exactly) = ADP + 3'-phosphoadenylyl sulfate

• Enzyme class 3: E.C.2.7.7.4 - Sulfate adenylyltransferase.
• Reaction: ATP + sulfate = diphosphate + adenylyl sulfate

ATP + sulfate = diphosphate + adenylyl sulfate (Bound ligand (Het Group name = ADX) corresponds exactly)

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

 Gene Ontology (GO) functional annotation 

• Biological process: sulfate assimilation (1 term)
• Biochemical function: transferase activity, transferring phosphorus-containing groups (3 terms)

reference

DOI no: 10.1016/j.jmb.2007.01.025

J Mol Biol 367:488-500 (2007)

PubMed id: 17276460

Elucidation of the active conformation of the APS-kinase domain of human PAPS synthetase 1.

N.Sekulic, K.Dietrich, I.Paarmann, S.Ort, M.Konrad, A.Lavie.

ABSTRACT

Bifunctional human PAPS synthetase (PAPSS) catalyzes, in a two-step process, the formation of the activated sulfate carrier 3'-phosphoadenosine 5'-phosphosulfate (PAPS). The first reaction involves the formation of the 5'-adenosine phosphosulfate (APS) intermediate from ATP and inorganic sulfate. APS is then further phosphorylated on its 3'-hydroxyl group by an additional ATP molecule to generate PAPS. The former reaction is catalyzed by the ATP-sulfurylase domain and the latter by the APS-kinase domain. Here, we report the structure of the APS-kinase domain of PAPSS isoform 1 (PAPSS1) representing the Michaelis complex with the products ADP-Mg and PAPS. This structure provides a rare glimpse of the active conformation of an enzyme catalyzing phosphoryl transfer without resorting to substrate analogs, inactivating mutations, or catalytically non-competent conditions. Our structure shows the interactions involved in the binding of the magnesium ion and PAPS, thereby revealing residues critical for catalysis. The essential magnesium ion is observed bridging the phosphate groups of the products. This function of the metal ion is made possible by the DGDN-loop changing its conformation from that previously reported, and identifies these loop residues unambiguously as a Walker B motif. Furthermore, the second aspartate residue of this motif is the likely candidate for initiating nucleophilic attack on the ATP gamma-phosphate group by abstracting the proton from the 3'-hydroxyl group of the substrate APS. We report the structure of the APS-kinase domain of human PAPSS1 in complex with two APS molecules, demonstrating the ability of the ATP/ADP-binding site to bind APS. Both structures reveal extended N termini that approach the active site of the neighboring monomer. Together, these results significantly increase our understandings of how catalysis is achieved by APS-kinase.

Selected figure(s)

Figure 1.
Figure 1. The APSK domain of human bifunctional PAPSS1 forms a symmetrical dimer and adopts the same overall fold as that observed in the context of the bifunctional enzyme. (a) Ribbon diagram of the APSK dimer (in light blue and orange) in complex with ADP and PAPS (maroon). The magnesium ion bridging the two nucleotides is depicted as a yellow sphere. Note that the N-terminal residues of one monomer approach the active site of the neighboring monomer. (b) Overlay of the APSK domain crystallized with ADP and PAPS (light blue), with APS (red), and with ADP and APS in the context of the full length PAPSS1 (gray, chain B in PDB entry 1XNJ). The rmsd for the two structures reported here is 0.44 Å over 201 common C^α atoms; between our structures and 1XNJ, it is 0.57–0.62 Å over 175–191 atoms. Important differences are: the conformation of the DGDN-loop (circle) and our ability to model nine additional residues at the N terminus (rectangle). These N-terminal residues approach the active site of the second monomer (the nucleotides of that monomer are included for orientation purposes). Figure 1. The APSK domain of human bifunctional PAPSS1 forms a symmetrical dimer and adopts the same overall fold as that observed in the context of the bifunctional enzyme. (a) Ribbon diagram of the APSK dimer (in light blue and orange) in complex with ADP and PAPS (maroon). The magnesium ion bridging the two nucleotides is depicted as a yellow sphere. Note that the N-terminal residues of one monomer approach the active site of the neighboring monomer. (b) Overlay of the APSK domain crystallized with ADP and PAPS (light blue), with APS (red), and with ADP and APS in the context of the full length PAPSS1 (gray, chain B in PDB entry 1XNJ). The rmsd for the two structures reported here is 0.44 Å over 201 common C^α atoms; between our structures and 1XNJ, it is 0.57–0.62 Å over 175–191 atoms. Important differences are: the conformation of the DGDN-loop (circle) and our ability to model nine additional residues at the N terminus (rectangle). These N-terminal residues approach the active site of the second monomer (the nucleotides of that monomer are included for orientation purposes).

Figure 4.
Figure 4. The DGDN-loop changes conformation in the presence of magnesium. (a) Stereo view of an overlay of the DGDN-loop (light blue) from the ADP–PAPS complex with the N terminus in orange and the previously reported ADP-APS complex (gray, PDB id 1XNJ). Due to the presence of magnesium in the ADP–PAPS complex, the DGDN-loop is stabilized at a conformation where Asp87 and Asp89 interact with a magnesium water ligand. This loop conformation, but not the DGDN-loop in the structure lacking magnesium, allows the N terminus to adopt the position seen in our structure. The three polar interactions between main chain atoms of the N terminus and the DGDN-loop are shown as dotted orange lines with the corresponding distances in Å. Pronounced differences in side-chain position are observed for Asp89 and Asn90 (black arrows). (b) Stereo view of the active site overlay between the APSK domain in complex with ADP plus PAPS (light blue), APS plus APS (red), and the monomer containing ADP plus APS in 1XNJ (gray). The presence of magnesium draws the DGDN-loop towards the active site, allowing Asp89 to interact with sugar hydroxyl groups (P)APS. In addition, the interaction between the PAPS 3′-phosphate group and the magnesium ion draws the nucleotide towards ADP. Figure 4. The DGDN-loop changes conformation in the presence of magnesium. (a) Stereo view of an overlay of the DGDN-loop (light blue) from the ADP–PAPS complex with the N terminus in orange and the previously reported ADP-APS complex (gray, PDB id 1XNJ). Due to the presence of magnesium in the ADP–PAPS complex, the DGDN-loop is stabilized at a conformation where Asp87 and Asp89 interact with a magnesium water ligand. This loop conformation, but not the DGDN-loop in the structure lacking magnesium, allows the N terminus to adopt the position seen in our structure. The three polar interactions between main chain atoms of the N terminus and the DGDN-loop are shown as dotted orange lines with the corresponding distances in Å. Pronounced differences in side-chain position are observed for Asp89 and Asn90 (black arrows). (b) Stereo view of the active site overlay between the APSK domain in complex with ADP plus PAPS (light blue), APS plus APS (red), and the monomer containing ADP plus APS in 1XNJ (gray). The presence of magnesium draws the DGDN-loop towards the active site, allowing Asp89 to interact with sugar hydroxyl groups (P)APS. In addition, the interaction between the PAPS 3′-phosphate group and the magnesium ion draws the nucleotide towards ADP.

The above figures are reprinted from an Open Access publication published by Elsevier: J Mol Biol (2007, 367, 488-500) copyright 2007.

Figures were selected by an automated process.