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Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaper › Journal article › Research › peer-review

Standard

Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state. / Aduri, Nanda G.; Ernst, Heidi A.; Prabhala, Bala K. et al.

In: Biochemical and Biophysical Research Communications, Vol. 495, No. 2, 08.01.2018, p. 1738-1743.

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaper › Journal article › Research › peer-review

Harvard

Aduri, NG, Ernst, HA, Prabhala, BK, Bhatt, S, Boesen, T, Gajhede, M & Mirza, O 2018, 'Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state', Biochemical and Biophysical Research Communications, vol. 495, no. 2, pp. 1738-1743. https://doi.org/10.1016/j.bbrc.2017.12.008

APA

Aduri, N. G., Ernst, H. A., Prabhala, B. K., Bhatt, S., Boesen, T., Gajhede, M., & Mirza, O. (2018). Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state. Biochemical and Biophysical Research Communications, 495(2), 1738-1743. https://doi.org/10.1016/j.bbrc.2017.12.008

CBE

Aduri NG, Ernst HA, Prabhala BK, Bhatt S, Boesen T, Gajhede M, Mirza O. 2018. Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state. Biochemical and Biophysical Research Communications. 495(2):1738-1743. https://doi.org/10.1016/j.bbrc.2017.12.008

MLA

Aduri, Nanda G. et al. "Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state". Biochemical and Biophysical Research Communications. 2018, 495(2). 1738-1743. https://doi.org/10.1016/j.bbrc.2017.12.008

Vancouver

Aduri NG, Ernst HA, Prabhala BK, Bhatt S, Boesen T, Gajhede M et al. Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state. Biochemical and Biophysical Research Communications. 2018 Jan 8;495(2):1738-1743. doi: 10.1016/j.bbrc.2017.12.008

Author

Aduri, Nanda G. ; Ernst, Heidi A. ; Prabhala, Bala K. et al. / Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state. In: Biochemical and Biophysical Research Communications. 2018 ; Vol. 495, No. 2. pp. 1738-1743.

Bibtex

@article{5952b0db796e4a0fa080c9045e45850a,
title = "Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state",
abstract = "The human proton coupled folic acid transporter PCFT is the major import route for dietary folates. Mutations in the gene encoding PCFT cause hereditary folic acid malabsorption, which manifests itself by compromised folate absorption from the intestine and also in impaired folate transport into the central nervous system. Since its recent discovery, PCFT has been the subject of numerous biochemical studies aiming at understanding its structure and mechanism. One major focus has been its oligomeric state, with some reports supporting oligomers and others a monomer. Here, we report the overexpression and purification of recombinant PCFT. Following detergent screening, n-Dodecyl beta-D-maltoside (DDM) and lauryl maltose neopentyl glycol (LMNG) were chosen for further work as they exhibited the most optimal solubilization. We found that purified detergent solubilized PCFT was able to bind folic acid, thus indicating a functionally active protein. Size exclusion chromatography showed that PCFT in DDM was polydisperse; the LMNG preparation was clearly monodisperse but with shorter retention time than the major DDM peak. To assess the oligomeric state negative stain electron microscopy was performed which showed a particle with the size of a PCFT dimer. (C) 2017 Elsevier Inc. All rights reserved.",
keywords = "Negative stain EM, Oligomeric state, PCFT, Size exclusion chromatography, Substrate binding assay, FOLATE TRANSPORTER, IMPACT, MEMBRANE, MALDI-TOFMS, CRYO-EM, EM STRUCTURE DETERMINATION, IDENTIFICATION, Proton-Coupled Folate Transporter/chemistry, Humans, Protein Multimerization, Spodoptera, Recombinant Proteins/chemistry, Sf9 Cells, Protein Structure, Quaternary, Glucosides, Solubility, Folic Acid/metabolism, Models, Molecular, Detergents, Microscopy, Electron, Animals, Ligands, Glycols",
author = "Aduri, {Nanda G.} and Ernst, {Heidi A.} and Prabhala, {Bala K.} and Shweta Bhatt and Thomas Boesen and Michael Gajhede and Osman Mirza",
year = "2018",
month = jan,
day = "8",
doi = "10.1016/j.bbrc.2017.12.008",
language = "English",
volume = "495",
pages = "1738--1743",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Elsevier Inc.",
number = "2",

}

RIS

TY - JOUR

T1 - Human proton coupled folic acid transporter is a monodisperse oligomer in the lauryl maltose neopentyl glycol solubilized state

AU - Aduri, Nanda G.

AU - Ernst, Heidi A.

AU - Prabhala, Bala K.

AU - Bhatt, Shweta

AU - Boesen, Thomas

AU - Gajhede, Michael

AU - Mirza, Osman

PY - 2018/1/8

Y1 - 2018/1/8

N2 - The human proton coupled folic acid transporter PCFT is the major import route for dietary folates. Mutations in the gene encoding PCFT cause hereditary folic acid malabsorption, which manifests itself by compromised folate absorption from the intestine and also in impaired folate transport into the central nervous system. Since its recent discovery, PCFT has been the subject of numerous biochemical studies aiming at understanding its structure and mechanism. One major focus has been its oligomeric state, with some reports supporting oligomers and others a monomer. Here, we report the overexpression and purification of recombinant PCFT. Following detergent screening, n-Dodecyl beta-D-maltoside (DDM) and lauryl maltose neopentyl glycol (LMNG) were chosen for further work as they exhibited the most optimal solubilization. We found that purified detergent solubilized PCFT was able to bind folic acid, thus indicating a functionally active protein. Size exclusion chromatography showed that PCFT in DDM was polydisperse; the LMNG preparation was clearly monodisperse but with shorter retention time than the major DDM peak. To assess the oligomeric state negative stain electron microscopy was performed which showed a particle with the size of a PCFT dimer. (C) 2017 Elsevier Inc. All rights reserved.

AB - The human proton coupled folic acid transporter PCFT is the major import route for dietary folates. Mutations in the gene encoding PCFT cause hereditary folic acid malabsorption, which manifests itself by compromised folate absorption from the intestine and also in impaired folate transport into the central nervous system. Since its recent discovery, PCFT has been the subject of numerous biochemical studies aiming at understanding its structure and mechanism. One major focus has been its oligomeric state, with some reports supporting oligomers and others a monomer. Here, we report the overexpression and purification of recombinant PCFT. Following detergent screening, n-Dodecyl beta-D-maltoside (DDM) and lauryl maltose neopentyl glycol (LMNG) were chosen for further work as they exhibited the most optimal solubilization. We found that purified detergent solubilized PCFT was able to bind folic acid, thus indicating a functionally active protein. Size exclusion chromatography showed that PCFT in DDM was polydisperse; the LMNG preparation was clearly monodisperse but with shorter retention time than the major DDM peak. To assess the oligomeric state negative stain electron microscopy was performed which showed a particle with the size of a PCFT dimer. (C) 2017 Elsevier Inc. All rights reserved.

KW - Negative stain EM

KW - Oligomeric state

KW - PCFT

KW - Size exclusion chromatography

KW - Substrate binding assay

KW - FOLATE TRANSPORTER

KW - IMPACT

KW - MEMBRANE

KW - MALDI-TOFMS

KW - CRYO-EM

KW - EM STRUCTURE DETERMINATION

KW - IDENTIFICATION

KW - Proton-Coupled Folate Transporter/chemistry

KW - Humans

KW - Protein Multimerization

KW - Spodoptera

KW - Recombinant Proteins/chemistry

KW - Sf9 Cells

KW - Protein Structure, Quaternary

KW - Glucosides

KW - Solubility

KW - Folic Acid/metabolism

KW - Models, Molecular

KW - Detergents

KW - Microscopy, Electron

KW - Animals

KW - Ligands

KW - Glycols

U2 - 10.1016/j.bbrc.2017.12.008

DO - 10.1016/j.bbrc.2017.12.008

M3 - Journal article

C2 - 29208467

AN - SCOPUS:85037609473

VL - 495

SP - 1738

EP - 1743

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 2

ER -