Crystallization and preliminary X-ray analysis of the vitamin D-binding protein from human serum

Koszelak, Stanley; McPherson, Alexander; Bouillon, Roger; Baelen, Hugo Van

doi:10.1016/0022-4731(85)90070-6

Cited by 6 publications

(2 citation statements)

References 5 publications

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“…While we present data that the quantifying peptides from VDBG are at a distance from the polymorphic amino acids in the protein and that they appear to be liberated at the same rate during trypsin proteolysis, it is not yet clear that the polymorphic peptides are released at the same rate. To evaluate the rate of peptide liberation in each haplotype and thus validate the ability to measure the concentration of each haplotype in plasma, purified VDBG protein from homozygotes will be required (37). …”

Section: Discussionmentioning

confidence: 99%

Measurement by a Novel LC-MS/MS Methodology Reveals Similar Serum Concentrations of Vitamin D–Binding Protein in Blacks and Whites

Henderson¹,

Lutsey²,

Misialek³

et al. 2016

Clinical Chemistry

129

107

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Background Vitamin D deficiency is associated with poor bone health and other adverse health outcomes. However, the associations are greatly attenuated in blacks compared with whites. One possible explanation for this attenuation could be different concentrations of bioavailable vitamin D metabolites in plasma, which are estimated using equations that include the total concentration of vitamin D binding globulin (VDBG) and haplotype-specific dissociation constants. Methods We developed a method to quantify VDBG using LC-MS/MS that could also identify the haplotypes/isoforms of VDBG present. We validated the method according to recent recommendations for publications of biomarker studies. We determined serum VDBG concentrations in samples from the Atherosclerosis Risk in Communities (ARIC) cohort and compared the results with a widely-used monoclonal immunoassay. Results An assay with a lower limit of quantification (<20% CV) of 71 µg/mL was developed using 10 µL of serum or plasma. The assay was linear from 64 to 434 µg/mL with total imprecision of 7.3–9.0% CV at ~250 µg/mL. Significant hemolysis interfered with quantification. The identification of isoforms was 97% concordant with genotyping (kappa coefficient). Method comparison with immunoassay revealed significant isoform-specific effects in the immunoassay. The concentration (SD) of VDBG by mass spectrometry was similar in whites and blacks [262 (25) vs. 266 (35) µg/mL, respectively; p=0.43]. Conclusions Validated mass spectrometric methods for the quantification of proteins in human samples are sensitive and specific and can provide additional information beyond immunoassay. Counter to prior observations by immunoassay, VDBG levels did not vary by race.

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Section: Discussionmentioning

confidence: 99%

Measurement by a Novel LC-MS/MS Methodology Reveals Similar Serum Concentrations of Vitamin D–Binding Protein in Blacks and Whites

Henderson¹,

Lutsey²,

Misialek³

et al. 2016

Clinical Chemistry

129

107

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“…(g) We have been working with vitamin D binding protein (VDBP) which is purified from pooled human plasma and comes to us in lyophilized form. This protein has been crystallized previously (Koszelak, McPherson, Bouillon & Van Baelen, 1985) but subsequent preparations have proved futile for crystal growth. Current samples of the protein form heavy brown precipitates or oils in virtually every precipitant we have investigated.…”

Section: Detergents As Additivesmentioning

confidence: 99%

Screening and optimization strategies for macromolecular crystal growth

Cudney¹,

Patel²,

Weisgraber³

et al. 1994

Acta Cryst D

258

230

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Today the determination of successful crystallization conditions for a particular macromolecule remains a highly empirical process. Sparse-matrix and grid-screening procedures are rapid and economical means to determine preliminary crystallization conditions. During optimization the variable set (pH, precipitant type and precipitant concentration) utilized in these procedures is screened in an attempt to determine appropriate conditions for the nucleation and growth of single crystals suitable for X-ray diffraction analysis. Unfortunately, in many cases this strategy will not produce single crystals suitable for X-ray diffraction analysis. We have explored, in an empirical sense, other tools for use during optimization. First, a new screening protocol is evaluated which employs less classical precipitating agents. Second, a set of 24 electrostatic crosslinking agents are evaluated for their ability to promote crystallization. Third, a panel of more than 30 detergents are evaluated for their ability to prevent sample aggregation and influence crystal growth.

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Crystallization and X-ray investigation of vitamin D-binding protein from human serum. Identification of the crystal content

Verboven¹,

Bondt²,

Ranter³

et al. 1995

The Journal of Steroid Biochemistry and Molecular Biology

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Crystallization and preliminary X-ray analysis of the vitamin D-binding protein from human serum

Cited by 6 publications

References 5 publications

Measurement by a Novel LC-MS/MS Methodology Reveals Similar Serum Concentrations of Vitamin D–Binding Protein in Blacks and Whites

Measurement by a Novel LC-MS/MS Methodology Reveals Similar Serum Concentrations of Vitamin D–Binding Protein in Blacks and Whites

Screening and optimization strategies for macromolecular crystal growth

Crystallization and X-ray investigation of vitamin D-binding protein from human serum. Identification of the crystal content

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