Natural Science Forum / Biology / Microbiology / March 2006

Signal peptides in E.coli using the sec system, as in any other gram
negative bacterium, follow very simple rules outlined in Gunnar von
Heijne's publications (they start with one or two positive residues,
then a hydrophobic stretch follows, and then the signal peptide ends
with two ala residues that is important for efficient peptide cleavage).
I'm quite confident that a signal peptide from Salmonella would work
efficiently in E.coli as long as it follows the rules for sec transport.

We have succesfully designed a new signal peptide based on these rules
which was able to translocate recombinant proteins very efficient*,
comparably to the commonly used signal peptides pelB and ompA. The
results showed that the efficiency of the signal peptide is gene
dependent, some signal peptides seem to work better with certain genes
than others. I don't believe that the difference seen in measurable
protein in the periplasm is necessarily caused by increased
translocation, but rather by an increase in expression, probably
translation.

*http://tinyurl.com/mscga

Trond Erik

You should check out the general secretory system.

the genes have names something like this...

secA, secB   etc...

The secretory sequence shouldn't be too hard to find.

If I recall correctly, there isn't a large diversity when it comes to
export signal sequence, so you probably won't find a "list", so much as
find a description of the motif which suggests probabilities of having
specific amino acid residues in certain positions relative to each other.

Then use PCR with custom primers to extract your gene of interest such
that the Sec signal peptide will be used to process your desired protein.

That is to say, if you can drop the signal sequence from s. enterica
which may not be processed correctly by DH5a, and replace it with a
correct one... you will be stylin.

You are going to have to retrieve the gene anyway, so custom primers
will be required... save yourself some time and headaches and do the
switch at the primer stage.

check out this paper

http://www.pubmedcentral.gov/articlerender.fcgi?tool=pubmed&pubmedid=12700260

it covers the sec and TAT pathways for excretion.

The sec pathway translocates proteins in an unfolded conformation, and
the TAT in a folded conformation.

This feature is useful for proteins which fold incorrectly in the periplasm.

pay particular attention to this section:

"Specificity of the precursor proteins for either the Sec pathway or the
Tat pathway seems to lie predominantly in the nature of the respective
signal peptides. Tat signal peptides possess a tripartite organization
similar to that of Sec signal peptides: a positively charged N terminus
(n-region) is followed by a core of hydrophobic amino acids (h-region)
and a more polar C terminus (c-region) preceding the signal peptidase
cleavage site (14, 30). In contrast to Sec signal sequences, Tat signal
peptides contain an (S/T)-R-R-X-F-L-K consensus motif (where X is highly
variable) at the boundary between the n- and the h-regions (1, 2). It is
reasonable to assume that one or more of the components of the Tat
pathway (but not of the Sec system) might recognize the precursor
proteins by the presence of this motif. However, additional features of
the signal peptides that go beyond the conserved sequence motif seem to
be important for successful recognition of the precursor proteins by
their respective translocase (3). Another important difference between
Tat and Sec signal peptides seems to reside in the hydrophobicity of
their h-regions, with Tat signal peptides being mostly less hydrophobic
than their Sec counterparts (9). Likewise, the hydrophobicity of the
n-region was reported to be another important factor determining Tat
pathway exclusivity (20)."