1: Biochim Biophys Acta. 2007 Dec 14 [Epub ahead of print]

Solution NMR of signal peptidase, a membrane protein.

Monika MS, Debra A K, Philip L Y.

Department of Molecular and Cell Biology, The University of Connecticut, Storrs, 
CT 06269, USA.

Useful solution nuclear magnetic resonance (NMR) data can be obtained from
full-length, enzymatically active type I signal peptidase (SPase I), an integral 
membrane protein, in detergent micelles. Signal peptidase has two transmembrane
segments, a short cytoplasmic loop, and a 27-kD C-terminal catalytic domain. It
is a critical component of protein transport systems, recognizing and cleaving
amino-terminal signal peptides from preproteins during the final stage of their
export. Its structure and interactions with the substrate are of considerable
interest, but no three-dimensional structure of the whole protein has been
reported. The structural analysis of intact membrane proteins has been
challenging and only recently has significant progress been achieved using NMR to
determine membrane protein structure. Here we employ NMR spectroscopy to study
the structure of the full-length SPase I in dodecylphosphocholine detergent
micelles. HSQC-TROSY spectra showed resonances corresponding to approximately 3/4
of the 324 residues in the protein. Some sequential assignments were obtained
from the 3D HNCACB, 3D HNCA, and 3D HN(CO) TROSY spectra of uniformly (2)H,
(13)C, (15)N-labeled full-length SPase I. The assigned residues suggest that the 
observed spectrum is dominated by resonances arising from extramembraneous
portions of the protein and that the transmembrane domain is largely absent from 
the spectra. Our work elucidates some of the challenges of solution NMR of large 
membrane proteins in detergent micelles as well as the future promise of these
kinds of studies.


PMID: 18177734 [PubMed - as supplied by publisher]

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