Zinc-free cubic pig insulin: Crystallization and structure determination

Dodson, E.J.; Dodson, Guy; Lewitova, Anita; Sabesan, M. N.

doi:10.1016/0022-2836(78)90409-6

Cited by 50 publications

(44 citation statements)

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“…X-ray data extending to 1.7 A resolution were processed (Oatley & French, 1982) to a final merging R factor of 0.07 (Dodson et al, 1978). Coordinates for molecule (1) of 2Zn insulin from which residues B1-B4, B29-B30 and side chains B5, B6 and BI0 had been removed were used in the calculation of rotation-translation functions.…”

Section: Previous Workmentioning

confidence: 99%

“…This paper reports the structure of dimeric insulin in the cubic crystal (space group /2~3 with a = 78.9 A) which is obtained in the absence of zinc at about pH 9. There are 24 molecules contained within the unit cell forming 12 exact (crystallographic) dimers (Dodson, Dodson, Lewitova & Sabesan, 1978). The symmetric dimer in this cubic crystal provides a standard for characterizing the departures from symmetry in the two molecules within the 2Zn insulin dimer.…”

Section: Introductionmentioning

confidence: 99%

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Structure of the pig insulin dimer in the cubic crystal

Badger¹,

Harris²,

Reynolds³

et al. 1991

Acta Crystallogr Sect B

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Atomic coordinates for pig insulin in the cubic crystal have been refined by reciprocal-space methods to an R factor of 0-173 for data between 10-0 and 1.7 A resolution with structure-factor amplitudes greater than two standard deviations. Stereochemical parameters for the refined model are close to standard values and the estimated error in the positions of well-ordered atoms is about 0.1 A. Residues directly involved in the formation of the exact (crystallographic) cubic insulin dimer are oriented similarly to * Current address:

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Section: Previous Workmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Structure of the pig insulin dimer in the cubic crystal

Badger¹,

Harris²,

Reynolds³

et al. 1991

Acta Crystallogr Sect B

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“…x-ray diffraction data extending from the 002 reflection to 1.7-A resolution has been measured (10), and atomic coordinates for the insulin monomer and 108 bound water molecules in the asymmetric unit were refined to an R factor of 0.20 (8). The 24 insulin molecules within the unit cell are arranged into rows parallel to the cube edges, leaving continuous interconnected solvent channels of diameter -30 A running through the crystal.…”

Section: Introductionmentioning

confidence: 99%

Water structure in cubic insulin crystals.

Badger

Caspar

1991

Proc. Natl. Acad. Sci. U.S.A.

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The electron density distribution of the solvent in the cubic insulin crystal structure, which occupies 65% of the volume, has been mapped from 1.7-A resolution diffraction data by an iterative difference Fourier method, using the previously determined protein structure as the refinement restraint. Starting with phases from the protein and a flat solvent model, the difference map calculated from the data was added outside the protein envelope, and the modified map was then used to recalculate phases for the iterative refinement. Tests of the method with model data, with the experimental data and a variant protein model, and by carrying out a partial refinement of the solvent map demonstrate that the refinement algorithm produces reliable values for the solvent density within the noise level of the data. Fluctuations in density are observed throughout the solvent space, demonstrating that nonrandom arrangements of the water molecules extend several layers from the well-ordered hydration shell in contact with the protein surface. Such ordering may account for the hydration force opposing close approach of hydrophilic surfaces and other long-range water-dependent interactions in living structures.x-ray diffraction data extending from the 002 reflection to 1.7-A resolution has been measured (10), and atomic coordinates for the insulin monomer and 108 bound water molecules in the asymmetric unit were refined to an R factor of 0.20 (8). The 24 insulin molecules within the unit cell are arranged into rows parallel to the cube edges, leaving continuous interconnected solvent channels of diameter -30 A running through the crystal. The 65% volume fraction of the crystal unit cell occupied by solvent corresponds to =440 water molecules per insulin molecule (Mr 5778). Thus, there is slightly more than one water molecule for each nonhydrogen protein atom. In addition, at the pH and ionic strength of the mother liquor, these crystals should contain about three Na' counterions and about one molecule of salt per insulin monomer. The low salt concentration and large solvent volume within these crystals provide a suitable system for examining the structure of water in the vicinity of a protein surface under near-physiological conditions. Our analysis of the crystal data demonstrates that the average distribution of water molecules beyond those in immediate contact with the protein surface is recognizably nonuniform.Water constitutes at least half of the volume in a wide variety of protein crystals (1), and much of this water is not uniquely ordered. Fourier transformation of the correctly phased Bragg diffraction amplitudes, measured by x-ray or neutron crystallography, maps the average distribution of scattering matter in the crystal asymmetric unit, regardless of the ordering of this matter. Phases for the Bragg reflections from well-ordered protein crystals can be calculated from atomic models (including ordered water) which have been refined (2, 3) to fit observed intensities within a small residual. (R factors for da...

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“…For the cubic insulin data, the overall merging residual, R m, was 0.071 for the 10 100 reflections profile fitted to 1.7A resolution (Dodson, Dodson, Lewitova & Sabesan, 1978):…”

Section: Tests and Practical Experience Of The Methodsmentioning

confidence: 99%