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Protein Science (2007), 16:2153-2165. Published by Cold Spring Harbor Laboratory Press. Copyright © 2007 The Protein Society
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Structural biology of transmembrane domains: Efficient production and characterization of transmembrane peptides by NMR

Jian Hu1,2,4, Huajun Qin1,4, Conggang Li1,2, Mukesh Sharma1,2, Timothy A. Cross1,2,3, and Fei Philip Gao1,2

1 Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306, USA
2 National High Magnetic Field Laboratory, Tallahassee, Florida 32310, USA
3 Institute of Molecular and Biophysics, Florida State University, Tallahassee, Florida 32306, USA

(RECEIVED May 10, 2007; FINAL REVISION June 20, 2007; ACCEPTED June 21, 2007)

Structural characterization of transmembrane peptides (TMPs) is justified because transmembrane domains of membrane proteins appear to often function independently of the rest of the protein. However, the challenge in obtaining milligrams of isotopically labeled TMPs to study these highly hydrophobic peptides by nuclear magnetic resonance (NMR) is significant. In the present work, a protocol is developed to produce, isotopically label, and purify TMPs in high yield as well as to initially characterize the TMPs with CD and both solution and solid-state NMR. Six TMPs from three integral membrane proteins, CorA, M2, and KdpF, were studied. CorA and KdpF are from Mycobacterium tuberculosis, while M2 is from influenza A virus. Several milligrams of each of these TMPs ranging from 25 to 89 residues were obtained per liter of M9 culture. The initial structural characterization results showed that these peptides were well folded in both detergent micelles and lipid bilayer preparations. The high yield, the simplicity of purification, and the convenient protocol represents a suitable approach for NMR studies and a starting point for characterizing the transmembrane domains of membrane proteins.

Keywords: membrane protein; transmembrane peptide; maltose binding protein; solution NMR; solid-state NMR


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J. Payandeh, C. Li, M. Ramjeesingh, E. Poduch, C. E. Bear, and E. F. Pai
Probing Structure-Function Relationships and Gating Mechanisms in the CorA Mg2+ Transport System
J. Biol. Chem., April 25, 2008; 283(17): 11721 - 11733.
[Abstract] [Full Text] [PDF]


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