Critical initial real-space refinement in the structure determination of arginine kinase

Genfa Zhou, Thayumanasamy Somasundaram, Eric Blanc, Zhi Chen, Michael Chapman

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Arginine kinase (AK), a homologue of creatine kinase, catalyses the reversible transfer of a phosphoryl group between a guanidino phosphate and ADP. The family of phosphagen kinases eluded structure determination for over 25 years until an inactive form creatine kinase (CK) structure was determined [Fritz-Wolf et al. (1996). Nature (London), 381, 341-345]. The structure determination of the active-form transition-state complex was non-trivial, owing to the distant relatedness and domain reorientation of AK compared with CK. Phases from a molecular-replacement solution of the large domain, supplemented by single isomorphous replacement and inter-crystal averaging, did not reveal interpretable electron density for the small domain. Reciprocal-space refinement of the initial model (R(free) = 0.54) by any of the commonly used methods, including post facto application of maximum-likelihood methods, led to overfitting without significant improvement of the partial initial model. By contrast, in the local real-space refinements which proved successful, the interdependence of atoms is limited to immediate neighbors, and atomic positions are not influenced by errors or omissions in remote parts of the structure. Modest improvement was possible without overfitting, and this was critical to the calculation of improved phases. Phases were refined and extended from 4.0 to 2.5 Å resolution by Fourier inversion of omit maps, combination with isomorphous replacement phases and averaging between crystal forms, after several batches of real- and reciprocal-space atomic refinement. The final structure refinement, against a 1.86 Å cryo data set yielded a high-quality model with R = 0.196 and R(free) = 0.224.

Original languageEnglish (US)
Pages (from-to)835-845
Number of pages11
JournalActa Crystallographica Section D: Biological Crystallography
Volume55
Issue number4
DOIs
StatePublished - Apr 1999
Externally publishedYes

Fingerprint

Arginine Kinase
creatine
Creatine Kinase
adenosine diphosphate
Crystals
wolves
Adenosine Diphosphate
Maximum likelihood
retraining
crystals
Carrier concentration
phosphates
Phosphotransferases
Phosphates
Electrons
inversions
Atoms
atoms

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biophysics
  • Condensed Matter Physics
  • Structural Biology

Cite this

Critical initial real-space refinement in the structure determination of arginine kinase. / Zhou, Genfa; Somasundaram, Thayumanasamy; Blanc, Eric; Chen, Zhi; Chapman, Michael.

In: Acta Crystallographica Section D: Biological Crystallography, Vol. 55, No. 4, 04.1999, p. 835-845.

Research output: Contribution to journalArticle

Zhou, Genfa ; Somasundaram, Thayumanasamy ; Blanc, Eric ; Chen, Zhi ; Chapman, Michael. / Critical initial real-space refinement in the structure determination of arginine kinase. In: Acta Crystallographica Section D: Biological Crystallography. 1999 ; Vol. 55, No. 4. pp. 835-845.
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abstract = "Arginine kinase (AK), a homologue of creatine kinase, catalyses the reversible transfer of a phosphoryl group between a guanidino phosphate and ADP. The family of phosphagen kinases eluded structure determination for over 25 years until an inactive form creatine kinase (CK) structure was determined [Fritz-Wolf et al. (1996). Nature (London), 381, 341-345]. The structure determination of the active-form transition-state complex was non-trivial, owing to the distant relatedness and domain reorientation of AK compared with CK. Phases from a molecular-replacement solution of the large domain, supplemented by single isomorphous replacement and inter-crystal averaging, did not reveal interpretable electron density for the small domain. Reciprocal-space refinement of the initial model (R(free) = 0.54) by any of the commonly used methods, including post facto application of maximum-likelihood methods, led to overfitting without significant improvement of the partial initial model. By contrast, in the local real-space refinements which proved successful, the interdependence of atoms is limited to immediate neighbors, and atomic positions are not influenced by errors or omissions in remote parts of the structure. Modest improvement was possible without overfitting, and this was critical to the calculation of improved phases. Phases were refined and extended from 4.0 to 2.5 {\AA} resolution by Fourier inversion of omit maps, combination with isomorphous replacement phases and averaging between crystal forms, after several batches of real- and reciprocal-space atomic refinement. The final structure refinement, against a 1.86 {\AA} cryo data set yielded a high-quality model with R = 0.196 and R(free) = 0.224.",
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