PDBsum entry 1lvc

Lyase

PDB id: 1lvc

Contents

Protein chains

485 a.a.

143 a.a. *

Ligands

DOT ×2

Metals

_YB ×3

_CA ×6

* Residue conservation analysis

PDB id:

1lvc

Name:

Lyase

Title:

Crystal structure of the adenylyl cyclase domain of anthrax edema factor (ef) in complex with calmodulin and 2' deoxy, 3' anthraniloyl atp

Structure:

Calmodulin-sensitive adenylate cyclase. Chain: a, b, c. Fragment: c-terminal domain (residues 291-800). Synonym: atp pyrophosphate-lyase, adenylyl cyclase, edema factor, ef, anthrax edema toxin adenylate cyclase component. Engineered: yes. Calmodulin. Chain: d, e, f. Engineered: yes

Source:

Bacillus anthracis. Organism_taxid: 1392. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693. Homo sapiens. Human. Organism_taxid: 9606.

Biol. unit:

Dimer (from PQS)

Resolution:

3.60Å R-factor: 0.281 R-free: 0.307

Authors:

Y.Shen,Y.-S.Lee,S.Soelaiman,P.Bergson,D.Lu,A.Chen,K.Beckingham, Z.Grabarek,M.Mrksich,W.-J.Tang

Key ref:

Y.Shen et al. (2002). Physiological calcium concentrations regulate calmodulin binding and catalysis of adenylyl cyclase exotoxins. EMBO J, 21, 6721-6732. PubMed id: 12485993 DOI: 10.1093/emboj/cdf681

Date:

28-May-02 Release date: 04-Dec-02

Protein chains

P40136  (CYAA_BACAN) -  Calmodulin-sensitive adenylate cyclase from Bacillus anthracis

Seq: 800 a.a.

Struc: 485 a.a.

Protein chains

P0DP23  (CALM1_HUMAN) -  Calmodulin-1 from Homo sapiens

Seq: 149 a.a.

Struc: 143 a.a.

Enzyme reactions

• Enzyme class: Chains A, B, C: E.C.4.6.1.1 - adenylate cyclase.
• Reaction: ATP = 3',5'-cyclic AMP + diphosphate

ATP (Bound ligand (Het Group name = DOT) matches with 75.00% similarity) = 3',5'-cyclic AMP + diphosphate

• Cofactor: Pyridoxal 5'-phosphate

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

reference

DOI no: 10.1093/emboj/cdf681

EMBO J 21:6721-6732 (2002)

PubMed id: 12485993

Physiological calcium concentrations regulate calmodulin binding and catalysis of adenylyl cyclase exotoxins.

Y.Shen, Y.S.Lee, S.Soelaiman, P.Bergson, D.Lu, A.Chen, K.Beckingham, Z.Grabarek, M.Mrksich, W.J.Tang.

ABSTRACT

Edema factor (EF) and CyaA are calmodulin (CaM)-activated adenylyl cyclase exotoxins involved in the pathogenesis of anthrax and whooping cough, respectively. Using spectroscopic, enzyme kinetic and surface plasmon resonance spectroscopy analyses, we show that low Ca(2+) concentrations increase the affinity of CaM for EF and CyaA causing their activation, but higher Ca(2+) concentrations directly inhibit catalysis. Both events occur in a physiologically relevant range of Ca(2+) concentrations. Despite the similarity in Ca(2+) sensitivity, EF and CyaA have substantial differences in CaM binding and activation. CyaA has 100-fold higher affinity for CaM than EF. CaM has N- and C-terminal globular domains, each binding two Ca(2+) ions. CyaA can be fully activated by CaM mutants with one defective C-terminal Ca(2+)-binding site or by either terminal domain of CaM while EF cannot. EF consists of a catalytic core and a helical domain, and both are required for CaM activation of EF. Mutations that decrease the interaction of the helical domain with the catalytic core create an enzyme with higher sensitivity to Ca(2+)-CaM activation. However, CyaA is fully activated by CaM without the domain corresponding to the helical domain of EF.

Selected figure(s)

Figure 4.
Figure 4 The activation of EF and CyaA-N by wild-type CaM and two series of CaM mutants. EF (1 nM) and CyaA-N (0.7 nM) were used for an adenylyl cyclase activity assay in the presence of 0.1 M free Ca^2+ with CaM mutants CaM 41/75 and 85/112. Each mutant has two cysteine mutations to lock either the N- or the C-terminal domain of CaM in the closed conformation (A and B). The same concentrations of EF and CyaA-N were used for an adenylyl cyclase activity assay in the presence of 1 M free Ca^2+ with CaM mutants B1Q, B2Q, B3Q and B4Q. Each mutant has a mutation inactivating one of four calcium-binding sites (C and D). Means SE are representative of at least two experiments.

Figure 5.
Figure 5 Effect of calcium and magnesium ions on adenylyl cyclase activity of EF and CyaA-N (A and B) and an EF mutant, EF-H577N (C and D). Adenylyl cyclase activity assays were performed with 10 M CaM at 0.1 M Ca^2+ (filled circles), 0.3 M Ca^2+ (open circles) and 1.0 M Ca^2+ (filled triangles) in the presence of 1 nM EF (A), and 0.7 nM CyaA-N (B). To analyze the mutant form of EF, adenylyl cyclase activities were measured with 10 M CaM and 1 nM EF or 66 nM EF-H577N with either 0.3 M Ca^2+ (C) or 10 mM Mg^2+ (D). Both were buffered by 10 mM EGTA. Maximal activities for EF in the magnesium titration are 1616 s^-1 (0.1 M Ca^2+), 1074 s^-1 (0.3 M Ca^2+), and 1009 s^-1 (1.0 M Ca^2+) (A) and those for CyaA-N are 2106 s^-1 (0.1 M Ca^2+), 1674 s^-1 (0.3 M Ca^2+) and 1385 s^-1 (1.0 M) (B). Maximal activities for EF and EF-H577N were 1208 s^-1 and 4 s^-1 (C) and those for EF and EF-H577N were 2726 s^-1 and 6 s^-1 (D), respectively. Means SE are representative of at least two experiments.

The above figures are reprinted from an Open Access publication published by Macmillan Publishers Ltd: EMBO J (2002, 21, 6721-6732) copyright 2002.

Figures were selected by an automated process.