Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1

Research output: Contribution to journal › Journal article › Research › peer-review

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Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1. / Rafiq, Maria; Ernst, Heidi A.; Aduri, Nanda G.; Prabhala, Bala K.; Tufail, Soban; Rahman, Moazur; Bloch, Magnus Borup; Mirza, Nadia; Taylor, Nicholas M.; Boesen, Thomas; Gajhede, Michael; Mirza, Osman.

In: Protein Expression and Purification, Vol. 190, 105990, 2022.

Research output: Contribution to journal › Journal article › Research › peer-review

Harvard

Rafiq, M, Ernst, HA, Aduri, NG, Prabhala, BK, Tufail, S, Rahman, M, Bloch, MB, Mirza, N, Taylor, NM, Boesen, T, Gajhede, M & Mirza, O 2022, 'Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1', Protein Expression and Purification, vol. 190, 105990. https://doi.org/10.1016/j.pep.2021.105990

APA

Rafiq, M., Ernst, H. A., Aduri, N. G., Prabhala, B. K., Tufail, S., Rahman, M., Bloch, M. B., Mirza, N., Taylor, N. M., Boesen, T., Gajhede, M., & Mirza, O. (2022). Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1. Protein Expression and Purification, 190, [105990]. https://doi.org/10.1016/j.pep.2021.105990

Vancouver

Rafiq M, Ernst HA, Aduri NG, Prabhala BK, Tufail S, Rahman M et al. Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1. Protein Expression and Purification. 2022;190. 105990. https://doi.org/10.1016/j.pep.2021.105990

Author

Rafiq, Maria ; Ernst, Heidi A. ; Aduri, Nanda G. ; Prabhala, Bala K. ; Tufail, Soban ; Rahman, Moazur ; Bloch, Magnus Borup ; Mirza, Nadia ; Taylor, Nicholas M. ; Boesen, Thomas ; Gajhede, Michael ; Mirza, Osman. / Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1. In: Protein Expression and Purification. 2022 ; Vol. 190.

Bibtex

@article{2f553767cc0746f28e8eef48d1ee7e9e,
title = "Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1",
abstract = "The human peptide transporter hPEPT1 (SLC15A1) is responsible for uptake of dietary di- and tripeptides and a number of drugs from the small intestine by utilizing the proton electrochemical gradient, and hence an important target for peptide-like drug design and drug delivery. hPEPT1 belongs to the ubiquitous major facilitator superfamily that all contain a 12TM core structure, with global conformational changes occurring during the transport cycle. Several bacterial homologues of these transporters have been characterized, providing valuable insight into the transport mechanism of this family. Here we report the overexpression and purification of recombinant hPEPT1 in a detergent-solubilized state. Thermostability profiling of hPEPT1 at different pH values revealed that hPEPT1 is more stable at pH 6 as compared to pH 7 and 8. Micro-scale thermophoresis (MST) confirmed that the purified hPEPT1 was able to bind di- and tripeptides respectively. To assess the insolution oligomeric state of hPEPT1, negative stain electron microscopy was performed, demonstrating a predominantly monomeric state.",
keywords = "Peptide transporter, hPEPT1, Protein expression, Protein purification, Binding studies, Negative-stain electron microscopy, BETA-LACTAM ANTIBIOTICS, PEPTIDE TRANSPORTERS, FAMILY SLC15, DRUG, RECOGNITION, INHIBITORS",
author = "Maria Rafiq and Ernst, {Heidi A.} and Aduri, {Nanda G.} and Prabhala, {Bala K.} and Soban Tufail and Moazur Rahman and Bloch, {Magnus Borup} and Nadia Mirza and Taylor, {Nicholas M.} and Thomas Boesen and Michael Gajhede and Osman Mirza",
year = "2022",
doi = "10.1016/j.pep.2021.105990",
language = "English",
volume = "190",
journal = "Protein Expression and Purification",
issn = "1046-5928",
publisher = "Academic Press",

}

RIS

TY - JOUR

T1 - Expression, purification and characterization of human proton-coupled oligopeptide transporter 1 hPEPT1

AU - Rafiq, Maria

AU - Ernst, Heidi A.

AU - Aduri, Nanda G.

AU - Prabhala, Bala K.

AU - Tufail, Soban

AU - Rahman, Moazur

AU - Bloch, Magnus Borup

AU - Mirza, Nadia

AU - Taylor, Nicholas M.

AU - Boesen, Thomas

AU - Gajhede, Michael

AU - Mirza, Osman

PY - 2022

Y1 - 2022

N2 - The human peptide transporter hPEPT1 (SLC15A1) is responsible for uptake of dietary di- and tripeptides and a number of drugs from the small intestine by utilizing the proton electrochemical gradient, and hence an important target for peptide-like drug design and drug delivery. hPEPT1 belongs to the ubiquitous major facilitator superfamily that all contain a 12TM core structure, with global conformational changes occurring during the transport cycle. Several bacterial homologues of these transporters have been characterized, providing valuable insight into the transport mechanism of this family. Here we report the overexpression and purification of recombinant hPEPT1 in a detergent-solubilized state. Thermostability profiling of hPEPT1 at different pH values revealed that hPEPT1 is more stable at pH 6 as compared to pH 7 and 8. Micro-scale thermophoresis (MST) confirmed that the purified hPEPT1 was able to bind di- and tripeptides respectively. To assess the insolution oligomeric state of hPEPT1, negative stain electron microscopy was performed, demonstrating a predominantly monomeric state.

AB - The human peptide transporter hPEPT1 (SLC15A1) is responsible for uptake of dietary di- and tripeptides and a number of drugs from the small intestine by utilizing the proton electrochemical gradient, and hence an important target for peptide-like drug design and drug delivery. hPEPT1 belongs to the ubiquitous major facilitator superfamily that all contain a 12TM core structure, with global conformational changes occurring during the transport cycle. Several bacterial homologues of these transporters have been characterized, providing valuable insight into the transport mechanism of this family. Here we report the overexpression and purification of recombinant hPEPT1 in a detergent-solubilized state. Thermostability profiling of hPEPT1 at different pH values revealed that hPEPT1 is more stable at pH 6 as compared to pH 7 and 8. Micro-scale thermophoresis (MST) confirmed that the purified hPEPT1 was able to bind di- and tripeptides respectively. To assess the insolution oligomeric state of hPEPT1, negative stain electron microscopy was performed, demonstrating a predominantly monomeric state.

KW - Peptide transporter

KW - hPEPT1

KW - Protein expression

KW - Protein purification

KW - Binding studies

KW - Negative-stain electron microscopy

KW - BETA-LACTAM ANTIBIOTICS

KW - PEPTIDE TRANSPORTERS

KW - FAMILY SLC15

KW - DRUG

KW - RECOGNITION

KW - INHIBITORS

U2 - 10.1016/j.pep.2021.105990

DO - 10.1016/j.pep.2021.105990

M3 - Journal article

C2 - 34637915

VL - 190

JO - Protein Expression and Purification

JF - Protein Expression and Purification

SN - 1046-5928

M1 - 105990

ER -

ID: 283758042