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#hydropathy plot

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Plasma Membrane Proteins

Which amino acid property is commonly analyzed in a hydropathy plot to predict transmembrane domains?

Computational prediction of membrane spans relies on physicochemical property hydrophobicity quantifying tendency to partition into non-polar phase. Kyte-Doolittle hydropathy scale derived from experimental water vapor transfer energies assigns isoleucine 4.5, valine 4.2, leucine 3.8, phenylalanine 2.8 as hydrophobic; arginine -4.5, lysine -3.9, asparagine -3.5, aspartate -3.5 as hydrophilic. Sliding window average length nineteen matches bilayer thickness smooths sequence revealing peaks sustained above cutoff indicating candidate transmembrane helices. Complementary scales include Goldman Engelman Steitz detecting helical hydrophobic moment and White Wimley interfacial octanol partitioning reflecting translocon insertion energetics. Algorithms combine hydrophobicity with charge distribution applying positive-inside rule where cytosolic loops enriched Arg Lys due to membrane potential, and with length constraints. Molecular weight does not correlate with membrane insertion, net charge alone insufficient due to loop variability, polarity ambiguous. Therefore hydrophobicity remains primary property analyzed in hydropathy plots for mapping membrane protein topology before high-resolution structures confirm assignments. Inclusion of positive-inside rule and signal peptide detection enhances prediction distinguishing signal anchors from true transmembrane helices for cloning strategies.

Ref: Kyte and Doolittle, Hydropathy Plot for Transmembrane Prediction, J Mol Biol 1982.

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Plasma Membrane Proteins

Which experimental technique is commonly used to determine the number of transmembrane domains in a protein?

Prediction of membrane topology begins with hydropathy profiling visualizing hydrophobic potential along polypeptide. Kyte-Doolittle scale assigns each amino acid numerical value reflecting water to vapor transfer free energy: isoleucine four point five, valine four point two, leucine three point eight highly hydrophobic, arginine minus four point five highly hydrophilic. Sliding window average typically nineteen to twenty one residues equivalent to hydrophobic thickness smooths profile revealing peaks above threshold about one point six indicating membrane-spanning segments. Positive-inside rule where arginine lysine enriched cytosolic loops aids orientation. Modern algorithms TMHMM Phobius TOPCONS integrate Hidden Markov Models with charge bias improving accuracy to about ninety percent. Experimental confirmation uses reporter fusions PhoA active periplasmic versus LacZ cytoplasmic, substituted cysteine accessibility method SCAM labeling membrane-impermeant reagents, and glycosylation mapping using acceptor sites inserted. SDS-PAGE separates by molecular weight, Western blotting detects immunoreactivity, FRAP measures lateral mobility but only hydropathy plot directly estimates number and positions of transmembrane hydrophobic segments guiding cloning and mutagenesis.

Ref: Kyte and Doolittle, A Simple Method for Displaying Hydropathic Character, J Mol Biol 1982.

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