PDBsum entry 3hpr

Transferase

PDB id: 3hpr

Contents

Protein chains

214 a.a. *

Ligands

AP5 ×2

Waters ×381

* Residue conservation analysis

PDB id:

3hpr

Name:

Transferase

Title:

Crystal structure of v148g adenylate kinase from e. Coli, in complex with ap5a

Structure:

Adenylate kinase. Chain: a, b. Synonym: ak, atp-amp transphosphorylase. Engineered: yes. Mutation: yes

Source:

Escherichia coli. Organism_taxid: 83333. Strain: k-12. Gene: adk, b0474, dnaw, jw0463, plsa. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.00Å R-factor: 0.210 R-free: 0.243

Authors:

V.J.Hilser,T.P.Travis,D.W.Bolen

Key ref:

T.P.Schrank et al. (2009). Rational modulation of conformational fluctuations in adenylate kinase reveals a local unfolding mechanism for allostery and functional adaptation in proteins. Proc Natl Acad Sci U S A, 106, 16984-16989. PubMed id: 19805185 DOI: 10.1073/pnas.0906510106

Date:

04-Jun-09 Release date: 03-Nov-09

Protein chains

P69441  (KAD_ECOLI) -  Adenylate kinase from Escherichia coli (strain K12)

Seq: 214 a.a.

Struc: 214 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.2.7.4.3 - adenylate kinase.
• Reaction: AMP + ATP = 2 ADP

AMP + ATP (Bound ligand (Het Group name = AP5) matches with 54.39% similarity) = 2 × ADP

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

reference

DOI no: 10.1073/pnas.0906510106

Proc Natl Acad Sci U S A 106:16984-16989 (2009)

PubMed id: 19805185

Rational modulation of conformational fluctuations in adenylate kinase reveals a local unfolding mechanism for allostery and functional adaptation in proteins.

T.P.Schrank, D.W.Bolen, V.J.Hilser.

ABSTRACT

Elucidating the complex interplay between protein structure and dynamics is a prerequisite to an understanding of both function and adaptation in proteins. Unfortunately, it has been difficult to experimentally decouple these effects because it is challenging to rationally design mutations that will either affect the structure but not the dynamics, or that will affect the dynamics but not the structure. Here we adopt a mutation approach that is based on a thermal adaptation strategy observed in nature, and we use it to study the binding interaction of Escherichia coli adenylate kinase (AK). We rationally design several single-site, surface-exposed glycine mutations to selectively perturb the excited state conformational repertoire, leaving the ground-state X-ray crystallographic structure unaffected. The results not only demonstrate that the conformational ensemble of AK is significantly populated by a locally unfolded state that is depopulated upon binding, but also that the excited-state conformational ensemble can be manipulated through mutation, independent of perturbations of the ground-state structures. The implications of these results are twofold. First, they indicate that it is possible to rationally design dynamic allosteric mutations, which do not propagate through a pathway of structural distortions connecting the mutated and the functional sites. Secondly and equally as important, the results reveal a general strategy for thermal adaptation that allows enzymes to modulate binding affinity by controlling the amount of local unfolding in the native-state ensemble. These findings open new avenues for rational protein design and fundamentally illuminate the role of local unfolding in function and adaptation.

Selected figure(s)

Figure 1.
Mutation strategy applied to adenylate kinase. Structure of “Open” [i.e., Apo-AK; (PDB ID 4AKE) (11)] and “Closed” [i.e., complex of AK and the nonhydrolysable bisubstrate analogue inhibitor P^1,P^5-Di(adenosine) pentaphosphate (Ap5A); PDB ID 1AKE (10)] states of AK. LID is shown in gray. The “LID domain,” as defined by Shapiro et al. (7). AMPbd is shown in green. The “AMP binding domain” (7). Red spheres, selected mutation sites.

Figure 4.
Surface Gly mutations in LID conserve ground-state structure. (A) Alignment of the reported crystal structures of WT and v148g AK. Shown in red are chains (WT and v148g) from position A within the asymmetric unit. Shown in black are chains from position B within the asymmetric unit. (B) ^1H-^15N HSQC spectra of WT (black) and v142g (red) AK at 21° C, which suppress local unfolding within the LID region. Enhanced contrast is used for v142g to allow visualization of peaks with decreased intensity, most likely because of exchange broadening. Available assignments are provided for resonances with differences at this temperature. (C) Analysis of structural perturbations effected by mutation. The gray spheres represent all atoms that move >0.3 Å from the WT to mutant structure in both copies within the ASU. The dark red spheres show the mutation site (position 148). The light red spheres show all perturbed atoms (gray) that can be connected to the mutation site by a continuous chain (< 6 Å per step) of other perturbed atoms. Blue spheres, Ap5A.

Figures were selected by an automated process.