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EpiQuest - H

EpiQuest-H allows plotting the hydrophobicity profile of a protein molecule using simoultaneously various matrixes.

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About EpiQuest-H

EpiQuest-H is a program that allows you to analyse the hydrophobicity of different regions in a protein sequence by simultaneously using several different algorithms for analysis (in contrast to standard programs of this kind that usually allow only one algorithm).

Matrix & Algorithm

Matrix & Algorithm

The same algorithm uses several different Matrices with amino-acid scale values. As some of the original scale values were prepared for hydrophobicity, and some for hydrophilic properties (all quite different in scale of the numbers), all of them were re-adjusted to approximately the same scale to provide comparable results from different matrices. The sources for the original ratios for amino acid hydrophobicity are the following:

# Matrix     Authors                                                             Reference

HKD2       Kyte J., Doolittle R.F.                                 J. Mol. Biol. 157:105-132(1982).

HWW2      White, S.H., and Wimley, W.C.                 Annual Review of Biophysics and Biomolecular Structure 28, 319–365. 

HBB2        Bull H.B., Breese K.                                   Arch. Biochem. Biophys. 161:665-670(1974).

HZZ2        Zhu, C., Gao, Y., Li, H. et al                       Proc Natl Acad Sci U S A 113, 12946–12951.

HHW2      Hopp T.P., Woods K.R.                              Proc. Natl. Acad. Sci. U.S.A. 78:3824-3828(1981).

HEW2       Eisenberg D., et al.                                   J. Mol. Biol. 179:125-142(1984).

HZL2        Zhao, G., London E.                                  Protein Sci. 15:1987-2001(2006).

Program output

Program Output

The program allows you to run all 7 different profiles in one go, each data presented as a histogram or as a bar indicates the areas that are hydrophilic according to the particular matrix used:

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hydro 1-epcam.gif

The image above shows the results of analysing for the hydrophobicity profile of EpCAM, a transmembrane molecule with a single transmembrane domain.You can see that the profile for this protein varies depending on the matrix used, although the TM domain is detected practically identically by all algorithms (the actual TM is marked by the red frame).When looking for the areas for a future antibody epitope we would advise to aim for single hydrophilic peak (in case this is a relevant issue for your analysis). 

Hydrophobicity and Acessibility

Hydrophobicity and Acessibility

It should be noted that Hydrophobicity does not correspond directly to Accessibility of the area at the surface of the folded protein molecule. Usually, the most hydrophilic areas are accessible (at the surface), while hydrophobic ones are not.

ep-cam ACC comparion with HY.gif

A hydrophobicity profile according to Kyte & Doolitle (above), when compared with accessibility profile (EpiQuest-A, below), shows that some hydrophilic areas are not necessarily the ones exposed at the surface of the protein, while some mildly hydrophobic may be accessible. Red frame indicates the transmembrane domain of the molecule, and while the C-terminal part of TM is not hydrophilic, it is likely fully accessible on a properly folded molecule.

Hydrophobicity and Antigenicity

Hydrophobicity and Antigenicity

There exists an old rule, in part originating from practical experience - if you are looking for a B-cell epitope to make an antibody, take the most hydrophilic fragment, and this will be a good epitope. With respect to accessibility of an epitope this is correct, however, there is no real correlation between the antigenic strength of an epitope and the sequence being hydrophilic.

hyspo-5 NS1 AG ACC HY relations.png

Above is a map of antigenic (red), accessible (blue) and hydrophilic (green. Matrix HRD2) for NS1 protein of Dengue virus 2. All main peaks of Antigenicity maps correspond to actual immunodominant epitopes of this protein. As you can see, the first strong epitope of NS1 is not hydrophilic, and the second largest peak is hydrophilic only in part. Here you also can see how relationships are between hydrophilic and accessible areas, which overlap only partially.

Analysis of Demo Sequences

For a full overview of the program's capabilities you can use Demo Mode. The Manual for EpiQuest-H explains how to and modify settings. You can also see the examples of how to analyse the sequences and interpret the results by visiting the Demo Sequences analysis page.

EpiQuest Suite and site www.epiquest.co.uk belongs to Aptum Biologics Ltd.

EpiQuest® is a registered Trademark of Aptum Biologics Ltd.

© 2018, 2020, 2021 Aptum Biologics Ltd.

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