Charge Distribution ofSignal Peptides and Signal Anchor Type II
Masahiro Gomi 1

gomi2@cc.tuat.ac.jp
12Takatsugu Hirokawa 12
takatsugu@din.or.jp

Shigeki Mitaku 1

mitaku@cc.tuat.ac.jp

Tokyo University of Agriculture and Technology, Department of Biotechnology, Tokyo
184-8588, Japan
Present address: Ryoka System Inc.

1 Introduction
Hydrophobic signal sequences, that is, signal peptides (SP) and type I and type II signal-anchor
sequences (SA-1, SA-2), are responsible for secretion of mature proteins and translocation of Cterminal and N-terminal segments, respectively [1]. Almost all signal sequences show positively charged
residues at their N-terminal end [2]. However, it is dicult to discriminate di erence between SP andSA from the information of amino acid sequences alone, because those sequences have very similar
characteristics [3]. Particularly, any clear sequence motif has not been reported so far in the C-terminal
end region at which SP is cut o . Only so-called 1-3 rule has been reported, which postulates that
small residues are usually observed at the position of 01 and 03 from the cleavage site of SP [3]. In
this work, we have carried out comparative studies of the charge distribution of SP and SA type IIand revealed signi cant helical periodicity of positive charge at the C-terminal end of SA type II.

2 Material and Methods
We have prepared three kinds of dataset from SWISS-PROT: (1) signal peptides from eukaryote, (2)the same type of sequences from prokaryote and (3) signal anchors type II. The datasets were selected
whose pair-wise sequence homology with any other sequences is lower than 25% and the number ofdata was 71 for SP, 123 for SA-II of eukaryote and 73 for SA-II of prokaryote. We have aligned all
sequences by the positively charged residues, K and R, ubiquitously found at the N-terminal end of
signal sequences. Then, we have calculated two parameters characterizing the charge distribution of
signal sequences: (1) net charges (the di erence between the number of positively charged residues
and negatively charged ones) per a supposed helix and (2) total charges (the sum of the number ofcharged residues) per a helix.

3 Results and Discussion
hen all amino acid sequences were aligned by a positively charged residue at the N-terminal end of
signal sequences, charged residues are rarely found in the hydrophobic regions from 1-st to about
15-th position. The lack of strongly polar residues in this region of SP and SA-2 is quite natural, sincepolar groups have large electric self energy in the nonpolar environment of membrane. Whereas, both
positive and negative charges increased at the C-terminal end. Although the distribution of charged
residues seems similar between SP and SA, the net charges and the total charges showed signi cantdi erence between the two kinds of sequences. The charge distribution in the region between 18-th
to 35-th residues exhibited marked helical periodicity of positive net charges for SA type II. However,amino acid sequences of SP in the same region did not show any helical periodicity and the average net363

charge was rather negative. This di erence in the distribution of net charges suggested that the helicalstructure of hydrophobic segments is extended to the C-terminal region in SA-II, and the putative
helices in C-terminal end region appear to interact with the negatively-charged part of machinery
for translocation of signal anchors. The distribution of total number of charged residues was similar
between SP and SA with an exception of the lack of charged residues at the positions 18-th and 19-th
residues in SA-II. This narrow and deep hole in the charge distribution could not be observed for SP ofeukaryote as well as prokaryote. Although the physical meaning of this feature is not clear yet, it may
be related to the cleavage of SP or the inhibition of the cleavage of SA. The di erence in the charge
distribution between SP and SA found in this work is statistically signi cant, and the discrimination
between the two types of signal sequences may become possible by analyzing the charge distribution
around the C-terminal end.

References[1] Plath, K., Mothes, W., Wilkinson, B.M., Stirling, C.J., Rapoport, T.A., Signal sequence recognition in posttranslational protein transport across the yeast ER membrane, Cell, 94(6):795{807,1998.
[2] Von Heijne, G., Computer-assisted identi cation of protein sorting signals and prediction of membrane proteins topology and structure, Advances in Computational Biol., 2:1{14, 1996.
[3] Nielsen, H., Engelbrecht, J., Brunak, S., and von Heijne, G., Identi cation of prokaryotic andeukaryotic signal prediction of their cleavage sites, Protein Eng., 10(1):1{6 , 1997.

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