Stability of Loops in the Structure of Lactose Permease

Abstract

Structural analysis of peptide fragments has provided useful information on the secondary structure of integral membrane proteins built from a helical bundle (up to seven transmembrane segments). Comparison of those results to recent X-ray crystallographic results showed agreement between the structures of the fragments and the structures of the intact proteins. Lactose permease of Escherichia coli (lac Y) offers an opportunity to test that hypothesis on a substantially larger integral membrane protein. Lac Y contains a bundle of 12 transmembrane segments connected by 11 loops. Eleven segments, each corresponding to one of the loops in this protein, were studied. Five of these segments form defined structures in solution as determined by multidimensional nuclear magnetic resonance. Four peptides form turns, and one peptide reveals the end of one of the transmembrane helices. These results suggest that some loops in helical bundles are stabilized by short-range interactions, particularly in smaller bundles, and such intrinsically stable loops may contribute to protein stability and influence the pathway of folding. Greater conformational flexibility may be found in large integral membrane proteins.

Biosynthesis and NMR-studies of a double transmembrane domain from the Y4 receptor, a human GPCR

“…A large body of literature supports the basic assumption of the model: For example, proteolysis of membrane proteins resulted in fragments containing entire TM sequences (Huang et al 1981), and chemically or recombinantly synthesized TM peptides spontaneously assembled thereby rescuing receptor activity (Kahn and Engelman 1992;Ridge et al 1995;Martin et al 1999;Wrubel et al 1994). Finally, peptides corresponding to the N and C terminus (Harmar 2001;O'Hara et al 1993), loop domains (Bennett et al 2004;Katragadda et al 2001a, b;Yeagle et al 2000) and transmembrane domains (Katragadda et al 2001a, b;Cohen et al 2008;Zheng et al 2006;Musial-Siwek et al 2008;Tian et al 2007;Lau et al 2008;Mobley et al 2007;Neumoin et al 2007) from GPCRs have been found to fold to distinct secondary structures which in certain cases resembled the structures of the corresponding regions of the intact receptor.…”

NMR structure of the transmembrane domain of the n-acetylcholine receptor β2 subunit

“…An understanding of how these factors affect ion channel function requires an understanding of how the transmembrane helices fold together to form a fully functional channel. The feasibility of a reductionist approach to understanding nAChR structure has been established by the crystal structure of the isolated EC domain of the mouse α1 nAChR subunit [15], and by comparing the crystal structures of other membrane proteins to NMR analyses of isolated domains [35, 36]. …”

Synergistic Interactions between Aqueous and Membrane Domains of a Designed Protein Determine its Fold and Stability

“…The contribution of loop domains to the overall structural stability, function and folding has been recognized for some membrane proteins. [11][12][13][14] To gain insights into the contributions of the extra-membranous domains to folding and stability of membrane proteins we have designed model peptides incorporating folding domains that span the membrane and extend into the aqueous phase. 15,16 As a model for this study, we have used MS1, a peptide system previously developed to elucidate the features influencing transmembrane helix association.…”