Proximity of helices VIII (Ala273) and IX (Met299) in the lactose permease of Escherichia coli

Qingda Wang; John C Voss; Wayne L. Hubbell; H. Ronald Kaback

doi:10.1021/bi972990f

Proximity of helices VIII (Ala273) and IX (Met299) in the lactose permease of Escherichia coli

Qingda Wang, John C Voss, Wayne L. Hubbell, H. Ronald Kaback

Biochemistry and Molecular Medicine

Research output: Contribution to journal › Article

19 Scopus citations

Abstract

Three double-Cys mutant pairs - Ala→273Cys/Met299→Cys, Thr266→Cys/Ile303→Cys, and Thr266→Cys/Ser306→Cys - were constructed in a functional lac permease construct devoid of Cys residues, and the excimer fluorescence or electron paramagnetic resonance (EPR) was studied with pyrene- or spin-labeled derivatives, respectively. After reconstitution into proteoliposomes, excimer fluorescence is observed with mutant Ala273→Cys/Met299→Cys, but not with the single-Cys mutants nor with mutants Thr266→Cys/Ile303→Cys or Thr266→Cys/Ser306→Cys. Furthermore, spin-spin interaction is also observed with mutant Ala273→Cys/Met299→Cys, but only after the permease is reconstituted into proteoliposomes. The results provide independent support for the conclusions that helix VIII is close to helix IX and that the transmembrane helices of the permease are more loosely packed in a detergent micelle as opposed to a phospholipid bilayer.

Original language	English (US)
Pages (from-to)	4910-4915
Number of pages	6
Journal	Biochemistry
Volume	37
Issue number	14
DOIs	https://doi.org/10.1021/bi972990f
State	Published - Apr 7 1998

ASJC Scopus subject areas

Biochemistry

Access to Document

10.1021/bi972990f

Fingerprint Dive into the research topics of 'Proximity of helices VIII (Ala273) and IX (Met299) in the lactose permease of Escherichia coli'. Together they form a unique fingerprint.

Cite this

Wang, Q., Voss, J. C., Hubbell, W. L., & Kaback, H. R. (1998). Proximity of helices VIII (Ala273) and IX (Met299) in the lactose permease of Escherichia coli. Biochemistry, 37(14), 4910-4915. https://doi.org/10.1021/bi972990f

@article{a0bbf93d87994d768ca1ea8d15e4bd03,

title = "Proximity of helices VIII (Ala273) and IX (Met299) in the lactose permease of Escherichia coli",

abstract = "Three double-Cys mutant pairs - Ala→273Cys/Met299→Cys, Thr266→Cys/Ile303→Cys, and Thr266→Cys/Ser306→Cys - were constructed in a functional lac permease construct devoid of Cys residues, and the excimer fluorescence or electron paramagnetic resonance (EPR) was studied with pyrene- or spin-labeled derivatives, respectively. After reconstitution into proteoliposomes, excimer fluorescence is observed with mutant Ala273→Cys/Met299→Cys, but not with the single-Cys mutants nor with mutants Thr266→Cys/Ile303→Cys or Thr266→Cys/Ser306→Cys. Furthermore, spin-spin interaction is also observed with mutant Ala273→Cys/Met299→Cys, but only after the permease is reconstituted into proteoliposomes. The results provide independent support for the conclusions that helix VIII is close to helix IX and that the transmembrane helices of the permease are more loosely packed in a detergent micelle as opposed to a phospholipid bilayer.",

author = "Qingda Wang and Voss, {John C} and Hubbell, {Wayne L.} and Kaback, {H. Ronald}",

year = "1998",

month = apr,

day = "7",

doi = "10.1021/bi972990f",

language = "English (US)",

volume = "37",

pages = "4910--4915",

journal = "Biochemistry",

issn = "0006-2960",

publisher = "American Chemical Society",

number = "14",

}

TY - JOUR

T1 - Proximity of helices VIII (Ala273) and IX (Met299) in the lactose permease of Escherichia coli

AU - Wang, Qingda

AU - Voss, John C

AU - Hubbell, Wayne L.

AU - Kaback, H. Ronald

PY - 1998/4/7

Y1 - 1998/4/7

N2 - Three double-Cys mutant pairs - Ala→273Cys/Met299→Cys, Thr266→Cys/Ile303→Cys, and Thr266→Cys/Ser306→Cys - were constructed in a functional lac permease construct devoid of Cys residues, and the excimer fluorescence or electron paramagnetic resonance (EPR) was studied with pyrene- or spin-labeled derivatives, respectively. After reconstitution into proteoliposomes, excimer fluorescence is observed with mutant Ala273→Cys/Met299→Cys, but not with the single-Cys mutants nor with mutants Thr266→Cys/Ile303→Cys or Thr266→Cys/Ser306→Cys. Furthermore, spin-spin interaction is also observed with mutant Ala273→Cys/Met299→Cys, but only after the permease is reconstituted into proteoliposomes. The results provide independent support for the conclusions that helix VIII is close to helix IX and that the transmembrane helices of the permease are more loosely packed in a detergent micelle as opposed to a phospholipid bilayer.

AB - Three double-Cys mutant pairs - Ala→273Cys/Met299→Cys, Thr266→Cys/Ile303→Cys, and Thr266→Cys/Ser306→Cys - were constructed in a functional lac permease construct devoid of Cys residues, and the excimer fluorescence or electron paramagnetic resonance (EPR) was studied with pyrene- or spin-labeled derivatives, respectively. After reconstitution into proteoliposomes, excimer fluorescence is observed with mutant Ala273→Cys/Met299→Cys, but not with the single-Cys mutants nor with mutants Thr266→Cys/Ile303→Cys or Thr266→Cys/Ser306→Cys. Furthermore, spin-spin interaction is also observed with mutant Ala273→Cys/Met299→Cys, but only after the permease is reconstituted into proteoliposomes. The results provide independent support for the conclusions that helix VIII is close to helix IX and that the transmembrane helices of the permease are more loosely packed in a detergent micelle as opposed to a phospholipid bilayer.

UR - http://www.scopus.com/inward/record.url?scp=0032492717&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032492717&partnerID=8YFLogxK

U2 - 10.1021/bi972990f

DO - 10.1021/bi972990f

M3 - Article

C2 - 9538009

AN - SCOPUS:0032492717

VL - 37

SP - 4910

EP - 4915

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 14

ER -