Q2.  How does the program determine transmembrane helices?

A2.  To idenify transmembrane helices, the SOSUI system (1) employs two
kinds of physical-chemical indices (an index by Kyte and Doolittle (2),
and a new amphiphilicity index proposed by us).	Predicted transmembrane
helices by SOSUI are classified into "primary" and "secondary"
classes.  We call this classification "a multi-mechanism hypothesis,
for formation of transmembrane helices", in which several physical-chemical
factors are important in secondary-structural formation of transmembrane
helices. 

  A "primary" transmembrane helix can be found out in the highly
hydrophobic, successive region of your query sequence by a hydropathy profile
based on the Kyte-Doolittle hydropathy index.  Such transmembrane helices
may consist mainly of apolar residues, and therefore may be inserted by
theirselves into the hydrophobic lipid-bilayer.  However, there will be less
hydrophobic transmembrane helices, which originate from some polar amino-acid
residues, than "primary" segments.  They must be detected for better
prediction of transmembrane helices.

  We defined an amphiphilicity index of polar amino acids, to predict
such less hydrophobic, "secondary" helices.  Polar residues are
often functionally significant in the interior of membrane proteins, for
example, in case of ion pairs of channel proteins.  Most of polar amino acids
have an amphiphilic part attached to a long hydrocarbon chain or a bulky
aromatic ring.  If a polar residue of a transmembrane helix locates near
the mebrane surface, its amphiphile would be contacted with water molecules
or polar head-groups of lipids for the thermodynamically stable state. 

  If a polar residue exists in the core of a transmembrane helix, it could
be explained in terms of the attractive polar interaction between transmembrane
helices (3), (4).  Our amphiphilicity index was determined as follows.

1)  We estimated phase-transfer energy of each amino acid, supposed by
       the accessible surface area of the hydrocarbon part.

2)  An amphiphilicity of each amino acid was calculated from the transfer
       energy above, by weighing with its intensity of polarity.

3)  Charged and non-charged polar residues were exemplified.

  In the SOSUI system, the profile based on the amiphiphilicity index
is employed to elongate the primary helices, and search for the secondary
ones.  A non-charged polar residue is favorable for the secondary segments
in our current predition.

  If a segment in your query is beyond the "primary" threshold
or the "secondary" threshold, it is predicted as a transmembrane
helix.  The current version of SOSUI puts an emphasis on descrimination of
transmembrane helices, but determination of an exact transmembrane region.
In the near future, the SOSUI system  will be also improved concerning these
points.
[References]

(1) Hirokawa, T., Seah, B-C., Mitaku, S.: Bioinformatics, 14 (4), 378-379 (1998)
(2) Kyte, J., Doolittle, R. F.: J. Mol. Biol., 157, 105-132 (1982)
(3) Mitaku, S., Suzuki, K., Odashima, S., Ikuta, K., Suwa, M., Kukita, F., Ishikawa, M., Itoh, H.: Proteins, 22, 350-362 (1995)
(4) Suwa, M., Hirokawa, T., Mitaku, S.: Proteins, 22, 363-377 (1995)