scholarly journals High-Throughout Simultaneous Detection of Vitamin B12 and Folate using Intrinsic Factor and Folate Binding Protein

2020 ◽  
pp. 5262-5276
Author(s):  
Chen Chen ◽  
Keyword(s):  
Vitamin B12 ◽  
Binding Protein ◽  
Intrinsic Factor ◽  
Simultaneous Detection ◽  
Folate Binding Protein

scholarly journals Ligand-induced conformation change in folate-binding protein

1993 ◽  
Vol 292 (3) ◽  
pp. 921-925 ◽  
Author(s):  
N C Kaarsholm ◽  
A M Kolstrup ◽  
S E Danielsen ◽  
J Holm ◽  
S I Hansen
Keyword(s):  
Fluorescence Spectroscopy ◽  
Ligand Binding ◽  
Binding Protein ◽  
Simultaneous Detection ◽  
Tryptophan Fluorescence ◽  
Random Coil ◽  
Folate Binding Protein ◽  
Beta Strand ◽  
Process Detection ◽  
Induced Aggregation

C.d. and fluorescence spectroscopy have been used to investigate the effect of ligand binding on the structure and stability of folate-binding protein (FBP) from cow's whey. The c.d. spectrum of unligated FBP predicts the following secondary structure: 22% helix, 25% antiparallel beta-strand, 5% parallel beta-strand, 17% turn and 31% random-coil structure. Folate binding to FBP results in significant changes in the c.d. spectrum. Analysis of the spectrum shows a 10% decrease in antiparallel beta-strand as a result of ligand binding. Folate binding also leads to strong quenching of FBP tryptophan fluorescence. The magnitude of the quench is proportional to ligand binding. The guanidinium chloride-induced unfolding of FBP is shown to be a multistate process. Detection by c.d. and fluorescence spectroscopy lead to non-identical transitions. Modelling studies are consistent with the existence of a stable folding intermediate. Ligand binding to FBP increases the apparent folding stability of the molecule. Simultaneous detection by c.d. and fluorescence indicate that the apparent increased folding stability is derived from ligand-induced aggregation of FBP.

Discordant Serum Folate Result Unique to the Beckman Access Assay.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3704-3704
Author(s):  
Lisa Wakeman ◽  
Roger Munro ◽  
John Lewis ◽  
Andrew Beddall ◽  
Saad Al-Ismail
Keyword(s):  
Vitamin B12 ◽  
Serum Ferritin ◽  
Intrinsic Factor ◽  
Reference Method ◽  
Calibration Method ◽  
Serum Folate ◽  
Vitamin Supplement ◽  
Folate Binding Protein ◽  
Endomysial Antibodies ◽  
Abbott Architect

Abstract Most current commercial assays for serum folate use folate-binding protein with chemiluminescence detection. Historically serum folate methods have been calibrated using pteroylglutamic acid (PGA) as this is more stable than 5Methyltetrahydrofolate (5MTHF) which is the form present in patients not receiving supplementation. Poor specificity of these binding assays together with different detection technologies and different method designs have highlighted the need for greater standardisation to calibration regimes so that individual folate species can be better identified. We describe here discrepant serum folate results in a patient which appear to be unrelated to the issues described above. Consequent to a procurement evaluation of multiple analytical systems, serum ferritin, vitamin B12 and folate assays were performed on the Beckman Access, Abbott Architect 12000SR, Bayer Centaur and Roche E170 analysers according to manufacturer’s instructions. Method comparison studies using serum samples from 500 individuals showed good agreement between all analytical systems for all three parameters. However, one serum sample from a 44 year old Caucasian male who had presented with weight loss, diarrhoea and a microcytic, hypochromic anaemia (Hb 10.7 g/L) gave discrepant serum folate results and was further investigated. He had no evidence of renal impairment. His only medication was a multivitamin tablet supplement which he had been taking for several weeks. IgA anti-TTG antibodies were 33 U/mL (NR 0 – 10) and IgA endomysial antibodies were weakly positive. There was a polyclonal increase in serum IgA = 4.62 g/L (NR 0.8 – 2.8). Otherwise routine chemistry, thyroid profile and a paraprotein screen were unremarkable. Parietal cell and intrinsic factor antibodies were negative. Serum ferritin = 16 ng/mL (NR 20 – 215); Vitamin B12 = 164 pg/mL (NR 150–600). The patient’s sample showed no discrepant inter-system results for ferritin or Vitamin B12. Serum folate assays (NR 3.0 – 15.0 ng/mL) were as follows: Technique Serum Folate Calibration method Beckman Access >20.0 PGA Roche E170 2.2 Quantaphase II Abbott Architect 1.9 Quantaphase II Bayer Centaur 1.5 5MTHF This pattern of results was confirmed on a second sample a month later. No discrepancies were found in twenty other patients with positive TTG and endomysial antibodies. Current methods for folate assays give different results and cannot measure the various forms of folate due to lack of calibrant standardisation with different types of automated analysers. In this patient, it may be that the folate species in the Vitamin supplement are measurable by the Beckman Access but are undermeasured with the other assay techniques. This data suggests the need for a reference method capable of identifying individual folate species in all patients. Technique Serum Folate Calibration method Beckman Access >20.0 PGA Roche E170 2.2 Quantaphase II Abbott Architect 1.9 Quantaphase II Bayer Centaur 1.5 5MTHF

The kidney in vitamin B12and folate homeostasis: characterization of receptors for tubular uptake of vitamins and carrier proteins

2006 ◽  
Vol 291 (1) ◽  
pp. F22-F36 ◽  
Author(s):  
Henrik Birn
Keyword(s):  
Intracellular Transport ◽  
Molecular Mechanisms ◽  
Binding Protein ◽  
Folate Receptor ◽  
Protein A ◽  
Intrinsic Factor ◽  
Vitamin B ◽  
Carrier Proteins ◽  
Folate Binding Protein ◽  
Renal Tubular

Over the past 10 years, animal studies have uncovered the molecular mechanisms for the renal tubular recovery of filtered vitamin and vitamin carrier proteins. Relatively few endocytic receptors are responsible for the proximal tubule uptake of a number of different vitamins, preventing urinary losses. In addition to vitamin conservation, tubular uptake by endocytosis is important to vitamin metabolism and homeostasis. The present review focuses on the receptors involved in renal tubular recovery of folate, vitamin B12, and their carrier proteins. The multiligand receptor megalin is important for the uptake and tubular accumulation of vitamin B12. During vitamin load, the kidney accumulates large amounts of free vitamin B12, suggesting a possible storage function. In addition, vitamin B12is metabolized in the kidney, suggesting a role in vitamin homeostasis. The folate receptor is important for the conservation of folate, mediating endocytosis of the vitamin. Interaction between the structurally closely related, soluble folate-binding protein and megalin suggests that megalin plays an additional role in the uptake of folate bound to filtered folate-binding protein. A third endocytic receptor, the intrinsic factor-B12receptor cubilin-amnionless complex, is essential to the renal tubular uptake of albumin, a carrier of folate. In conclusion, uptake is mediated by interaction with specific endocytic receptors also involved in the renal uptake of other vitamins and vitamin carriers. Little is known about the mechanisms regulating intracellular transport and release of vitamins, and whereas tubular uptake is a constitutive process, this may be regulated, e.g., by vitamin status.

Cubilin, the Intrinsic Factor-Vitamin B12 Receptor in Development and Disease

2020 ◽  
Vol 27 (19) ◽  
pp. 3123-3150 ◽  
Author(s):  
Renata Kozyraki ◽  
Olivier Cases
Keyword(s):  
Vitamin B12 ◽  
Cancer Progression ◽  
Intrinsic Factor ◽  
Basic Research ◽  
Toxic Substances ◽  
Human Pathology ◽  
Peripheral Membrane Protein ◽  
Epidermal Growth ◽  
Fgf8 Signaling

Gp280/Intrinsic factor-vitamin B12 receptor/Cubilin (CUBN) is a large endocytic receptor serving multiple functions in vitamin B12 homeostasis, renal reabsorption of protein or toxic substances including albumin, vitamin D-binding protein or cadmium. Cubilin is a peripheral membrane protein consisting of 8 Epidermal Growth Factor (EGF)-like repeats and 27 CUB (defined as Complement C1r/C1s, Uegf, BMP1) domains. This structurally unique protein interacts with at least two molecular partners, Amnionless (AMN) and Lrp2/Megalin. AMN is involved in appropriate plasma membrane transport of Cubilin whereas Lrp2 is essential for efficient internalization of Cubilin and its ligands. Observations gleaned from animal models with Cubn deficiency or human diseases demonstrate the importance of this protein. In this review addressed to basic research and medical scientists, we summarize currently available data on Cubilin and its implication in renal and intestinal biology. We also discuss the role of Cubilin as a modulator of Fgf8 signaling during embryonic development and propose that the Cubilin-Fgf8 interaction may be relevant in human pathology, including in cancer progression, heart or neural tube defects. We finally provide experimental elements suggesting that some aspects of Cubilin physiology might be relevant in drug design.

scholarly journals Increased vitamin D binding protein levels are associated with irritable bowel syndrome

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Elif Börekci ◽  
Mahmut Kılıç ◽  
Zeynep Ozan ◽  
Hasan Börekci ◽  
Tekin Yıldırım ◽  
...  
Keyword(s):  
Vitamin D ◽  
Logistic Regression ◽  
Irritable Bowel Syndrome ◽  
Regression Analysis ◽  
Vitamin B12 ◽  
Logistic Regression Analysis ◽  
Binding Protein ◽  
Serum Levels ◽  
Vitamin D Binding Protein ◽  
Irritable Bowel

Abstract Objectives There is no reliable and valid biomarker to identify Irritable bowel syndrome (IBS) and its subtypes. The aim of this study is to explore potential serum biomarkers that may be associated with IBS subtypes, particularly in the vitamin D pathway. Methods The study population comprised 75 IBS patients and 79 controls. Patients divided into IBS subtypes. Routine biochemical parameters, 25-OH-vitamin D, vitamin D binding protein (VDBP) and vitamin D receptor (VDR) serum levels were compared between IBS subtypes and controls. Factors related to IBS subtypes were examined by multivariate logistic regression analysis. Results Vitamin D levels were lower; VDBP and VDR were higher in all IBS patients than in controls (p<0.001; 0.047 and 0.029, respectively). According to logistic regression analysis, VDBP was a disease-related parameter as much as vitamin D in all IBS subtypes. C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were higher especially in diarrhea-dominant IBS (IBS-D) (p=0.041; 0.046) and vitamin B12 were significantly lower in constipation-dominant IBS (IBS-C) (p=0.001). Conclusions Increased VDBP levels were associated with all IBS subtypes. Patients, especially in IBS-D, had higher serum levels of VDBP, CRP and ESR. Vitamin B12 deficiency, which we consider as a result of the disease, was more common in IBS-C.

Divalent cations and vitamin B12 uptake by intestinal brush-border membrane vesicles

1980 ◽  
Vol 239 (6) ◽  
pp. G452-G456
Author(s):  
R. C. Beesley ◽  
C. D. Bacheller
Keyword(s):  
Vitamin B12 ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Divalent Cations ◽  
Intrinsic Factor ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Tetraacetic Acid ◽  
Intestinal Brush Border Membrane ◽  
Intestinal Brush Border

Brush-border membrane vesicles from hamster intestine were employed to investigate uptake (binding) of vitamin B12 (B12). Ileal vesicles took up 25 times more B12 than did jejunal vesicles. Uptake of B12 by ileal vesicles was dependent on intrinsic factor (IF) and required Ca2+. Increasing the Ca2+ concentration caused an increase in uptake of B12 reaching a maximum at approximately 8 mM Ca2+. At high Ca2+ concentrations, 6–8 mM, Mg2+ had little effect on uptake of B12. At low Ca2+ concentrations, up to 2 mM, Mg2+ stimulated B12 uptake. Mg2+, Mn2+, and, to a lesser extent, Sr2+ stimulated Ca2+-dependent B12 uptake, but Zn2+, Ba2+, Na+, K+, and La3+ did not. B12 was apparently not metabolized and was bound as IF-B12 complex, which could be removed with (ethylenedinitrilo)tetraacetic acid (EDTA). Our results suggest that two types of divalent cation reactive sites are involved in binding of IF-B12. One is Ca2+ specific. The other is less specific reacting with Mg2+, Mn2+, Sr2+, and perhaps Ca2+ itself, thereby stimulating Ca2+-dependent binding of IF-B12 to its ileal receptor.

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