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#protein structure

114 public questions tagged with this topic.

Plasma Membrane Proteins

Which of the following is a common feature of transmembrane β-barrel proteins?

Beta-barrel membrane proteins display characteristic functional and structural attributes correlating with aqueous pore formation. Architecture comprises even number amphipathic beta-strands eight to twenty four organized antiparallel hydrogen bonded sheet closed cylindrically first strand bonded last strand strands tilted thirty to sixty degrees relative to barrel axis. Even residues hydrophobic facing lipid core, odd hydrophilic facing lumen generating water-filled channel diameter about seven to fifteen angstroms variable. Extracellular loops often long folding into pore constriction loop three in OmpF forming eyelet governing size exclusion approximately six hundred daltons cutoff and charge selectivity via acidic glutamate aspartate and basic arginine lysine lining. These porins facilitate passive diffusion of small polar nutrients including sugars, amino acids, phosphate, nucleosides, and antibiotics down concentration gradient rates up to million per second without energy, essential for Gram-negative bacterial survival. Trimeric assembly stabilizes. Eukaryotic mitochondria VDAC transports ATP ADP metabolites. Alpha-helical transporters active secondary carriers elsewhere, beta-barrels exclusively allow diffusion of small polar molecules through relatively nonselective aqueous pore.

Ref: Nikaido, Molecular Basis of Bacterial Outer Membrane Permeability, Microbiol Mol Biol Rev 2003.

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Lipid Link Protein

What is the length of a single transmembrane α-helix in lipid bilayers if it spans ~3 nm?

Geometrical constraints of fluid phospholipid bilayer dictate minimal span length for alpha-helical membrane proteins. Hydrophobic thickness between carbonyl regions of liquid-crystalline phosphatidylcholine bilayer averages three point zero nanometers. An alpha helix advances zero point fifteen nanometers per residue along axis with three point six residues per turn, pitch zero point five four nanometers stabilized by intramolecular hydrogen bonds between carbonyl i and amide i plus four. Dividing three nanometers by zero point fifteen per residue yields about twenty residues required for perpendicular crossing without exposure of polar backbone. Surveys of high-resolution structures and hydropathy analyses reveal hydrophobic stretches of nineteen to twenty three residues predominantly leucine, isoleucine, valine and phenylalanine flanked by interfacial aromatic belt tryptophan tyrosine at glycerol region and positively charged lysine arginine following positive-inside rule determining orientation. Glycophorin A helix of nineteen residues exemplifies minimal length with dimerization via GXXXG motif. Longer helices tilt up to thirty degrees accommodating mismatch or contain proline kinks influencing channel gating and receptor activation.

Ref: Alberts et al., Molecular Biology of the Cell, 6th ed., Chapter 10, Transmembrane Alpha-Helix Length and Bilayer Thickness.

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Lipid Link Protein

Which detergent is most likely to maintain a protein’s native structure during extraction?

Extraction of integral membrane proteins for functional study demands detergent that disrupts lipid bilayer but avoids complete unfolding of polypeptide. Ionic strong detergents like SDS bind cooperatively to protein backbone at about one SDS per two amino acids, imparting negative charge, disrupting secondary structure and denaturing enzymes, suitable for electrophoresis but not activity assays. Non-ionic detergents Triton X-100, Nonidet P-40, dodecyl-beta-D-maltoside DDM and octyl glucoside are milder; they replace lipids around transmembrane helices forming protein-detergent micelles, maintaining helical packing and often preserving oligomerization and cofactor binding. Among options list, Triton X-100 is mildest retaining native structure of many receptors, transporters and photosynthetic complexes, though still can inactivate sensitive proteins. Sodium deoxycholate is bile salt anionic more denaturing, disrupting protein-protein contacts. Selection depends on protein stability: mild non-ionic for functional reconstitution, harsher ionic for size analysis. Thus Triton X-100 is preferred when aim is to maintain native conformation during solubilization for downstream assays and crystallization trials.

Ref: Privé, Methods 2007, Detergent choice; Garavito & Ferguson-Miller, J Biol Chem 2001.

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Cytoskeleton

Which of the following proteins is a component of intermediate filaments?

Intermediate filament protein family comprises over 70 members classified by sequence homology and expression pattern regulated developmentally. Signature central rod domain about 310 amino acids alpha helical coiled coil flanked by variable head tail domains mediating assembly and interactions. Major subgroups include type I acidic keratins 28 genes and type II basic keratins 26 genes obligate heteropolymers in epithelia forming extensive cytoskeleton; type III includes vimentin mesenchymal fibroblasts endothelial desmin skeletal cardiac smooth muscle GFAP astrocytes peripherin peripheral neurons; type IV neurofilaments NF light medium heavy in central neurons determining axon caliber; type V lamins nuclear envelope; type VI nestin progenitor cells. Keratin represents classic component forming dense tonofilament networks resisting mechanical stress chemical insult and apoptosis. Tubulin alpha beta heterodimers make microtubules actin forms thin filaments myosin motor domain binds actin not intermediate. Therefore keratin is recognized intermediate filament constituent essential for epithelial integrity mutations causing epidermolysis bullosa simplex due to cell fragility under friction trauma.

Ref: Eriksson et al., Physiol Rev 2009 – Intermediate filament proteins including keratin as component classification.

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Cytoskeleton

The stalk domain of kinesin-1 is responsible for:

Domain architecture of kinesin-1 reveals clear division of labor essential for processive dimer motor function. N terminal globular motor domain about 340 amino acids contains P loop ATPase site switch I II microtubule binding interface and neck linker for mechanochemistry converting ATP hydrolysis to mechanical step. Immediately C terminal lies neck coiled coil ensuring dimerization of motor domains upon folding critical for hand over hand coordination followed by elongated stalk extending about 50 nm formed by interrupted coiled coil heptad repeats left handed supercoils stabilizing heavy chain dimer through hydrophobic packing of a and d positions leucine zipper like. Disruption of stalk by mutation prevents dimer formation and abolishes processivity yielding monomeric non processive motor. Central hinge regions provide flexibility for autoinhibited folding where C terminal tail interacts with motor domains to block ATPase until cargo binding unfolds. C terminal globular tail binds kinesin light chains via heptad interactions adapting to diverse vesicles mitochondria mRNA. Thus stalk responsible for dimerization via coiled coil interaction not ATP hydrolysis.

Ref: Hirokawa & Noda, Physiol Rev 2008 – Kinesin-1 stalk coiled coil mediated dimerization domain function.

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Protein Sorting and Trafficking

Type I membrane proteins have:

Single-pass membrane proteins classified by orientation and presence of cleavable signal define major group of surface receptors. Type I proteins possess cleavable N-terminal signal sequence of fifteen to thirty residues that targets ribosome to Sec61 and removed co-translationally by signal peptidase complex, leaving new N-terminus in ER lumen later becoming extracellular after trafficking. Downstream hydrophobic stretch acts as stop-transfer anchor halting translocation and spanning bilayer once with orientation N-lumenal, C-cytosolic during insertion, corresponding to N-exoplasmic, C-cytosolic at plasma membrane. Many immune receptors, growth factor receptors, viral glycoproteins and type I cytokines follow this blueprint; ectodomain often glycosylated and ligand-binding, cytosolic tail containing signaling motifs phosphorylated by kinases. Type II proteins use uncleaved signal-anchor with opposite N-cytosolic C-lumenal orientation, Type III also uncleaved but reversed charge distribution giving N-lumenal orientation. No cleavable C-terminal signal typical. Distinguishing cleavage pattern explains domain exposure, glycosylation topology and accessibility for drug targeting and antibody recognition, important for therapeutic design and understanding receptor activation mechanisms. Additional coordination with cellular stress pathways ensures fidelity, prevents aggregation, and links trafficking to growth control and proteostasis maintenance across diverse cell types and developmental stages.

Ref: Spiess & Beuret, Curr Opin Cell Biol 11: 1999, Type I Membrane Protein Topology.

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