We have observed a common sequence motif in membrane proteins, which we call a glycine zipper. Glycine zipper motifs are strongly overrepresented and conserved in membrane protein sequences, and mutations in glycine zipper motifs are deleterious to function in many cases. The glycine zipper has a significant structural impact, engendering a strong driving force for right-handed packing against a neighboring helix. Thus, the presence of a glycine zipper motif leads directly to testable structural hypotheses, particularly for a subclass of glycine zipper proteins that form channels. For example, we suggest that the membrane pores formed by the amyloid-␤ peptide in vitro are constructed by glycine zipper packing and find that mutations in the glycine zipper motif block channel formation. Our findings highlight an important structural motif in a wide variety of normal and pathological processes.amyloid-␤ ͉ membrane channel ͉ membrane protein structure ͉ prion ͉ transmembrane helix M ore than 13 structures a day are currently being deposited in the Protein Data Bank (1), and structural genomics centers have been created to obtain structures even faster [such as the National Institute of General Medical Sciences (NIGMS) Protein Structure Initiative, www.nigms.nih.gov͞psi]. In this assault on protein structure, however, technical challenges have left membrane proteins far behind. Membrane protein structures are currently being solved at Ϸ0.2% of the pace of soluble proteins (2). Thus, membrane protein biochemists are relatively starved for structural insight into these key proteins. In the absence of dramatic technical improvements, alternatives to experimental structure determination are needed.Here, we describe a transmembrane (TM) sequence motif, the glycine zipper, that can lead directly to structural models for many membrane proteins. The most significant glycine zipper sequence patterns are (G,A,S)XXXGXXXG and GXXXGX-XX(G,S,T). These patterns contain a GXXXG motif, which is known to be important in TM helix homodimers where the Gly faces are in direct contact (3-5). The GXXXG sequence pattern is statistically overrepresented in membrane proteins in general, not just in TM homodimers (4). Nevertheless, the structural role of the GXXXG pattern in other types of TM helix packing interactions has not been elucidated. We find that the addition of an appropriately spaced small residue, as found in the glycine zipper, leads to a distinct preference for right-handed packing against a heterologous helix surface. Thus, the presence of a glycine zipper generates a strong helix packing prediction, particularly for homooligomeric channel proteins, providing a structural foundation for hypothesis-driven investigations. MethodsGlycine Zipper Motif Search. We started with Swiss-Prot release 41.15 containing 129,996 proteins (6). All sequences Ͻ50 residues in length were removed, leaving 125,887 proteins. Helical membrane proteins were identified by using the Eisenberg hydrophobicity scale and a window length of 21 residues (7). ...

Current procedures for manual extraction of mature muscle tissue in micromechanical structures are time consuming and can damage the living components. To overcome these limitations, we have devised a new system for assembling muscle-powered microdevices based on judicious manipulations of materials phases and interfaces. In this system, individual cells grow and self-assemble into muscle bundles that are integrated with micromechanical structures and can be controllably released to enable free movement. Having realized such an assembly with cardiomyocytes we demonstrate two potential applications: a force transducer able to characterize in situ the mechanical properties of muscle and a self-assembled hybrid (biotic/abiotic) microdevice that moves as a consequence of collective cooperative contraction of muscle bundles. Because the fabrication of silicon microdevices is independent of the subsequent assembly of muscle cells, this system is highly versatile and may lead to the integration of cells and tissues with a variety of other microstructures.

Although membrane channel proteins are important to drug discovery and hold great promise as engineered nanopore sensing elements, their widespread application to these areas has been limited by difficulties in fabricating planar lipid-bilayer membranes. We present a method for forming these sub-5-nm-thick free-standing structures based on a self-assembly process driven by solvent extraction in a microfluidic channel. This facile automatable process forms high-quality membranes able to host channel proteins measurable at single-molecule conductance resolution.

Nanopores have been explored as highly sensitive sensors for detection and rapid sequencing of single molecules of DNA. To sequence DNA with a nanopore requires that adenine (A), cytosine (C), thymine (T), and guanine (G) produce distinct current signals as they traverse the pore. Recently, we demonstrated that homopolymers of adenine, cytosine, and thymine immobilized in the nanopore protein alpha-hemolysin (alphaHL) produced distinct current blockades dependent on their chemical orientation. To probe the detection limit of alphaHL, we examined immobilized single strands of T(40) DNA (polyT) with single base substitutions of A, C, and G at 12 positions on the strand occupying the stem region of alphaHL. We find blockade currents sensitive to base identity over most of these positions with the most sensitive region near the pore constriction. Adenine substitutions increase the measured blockade current to values intermediate to the polyT and polyA currents at a number of positions, while C substitutions increase the current to a level intermediate to polyT and polyC values in some positions, but decrease it below polyT in others. These changes in blockade current were also observed for G substitutions. These results indicate that total blockade currents measured in alphaHL arise from nucleotides at multiple locations and thus are not uniquely attributable to an individual base in a specific position, a finding consistent with a recently published study. The measurements of C and G substitutions also suggest that blockade current may be modulated through interactions between nucleotides and the pore interior at multiple sites in alphaHL.