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Hydrophobic Membrane Translocating Sequence Peptides 127
TABLE 6.2
Commonly Used Translocating Peptides
Name Sequence
HIV Tat peptide
Penetratin
Penetratin analogue
Transportan
SV40 large T antigen NLS (monopartite)
Nucleoplasmin NLS (bipartite)
kFGF MTS
kFGF MTS analogue
such peptide. One advantage of this approach is the use of fully free peptides and
proteins, which obviates the need for chemical modification that might affect biological
activity. As well as being synthetically straightforward and compatible with
existing solid-phase peptide synthesis methods, the thiazolidine formation reaction
is site-specific and avoids potential interference with the biological activity of the
cargo. Furthermore, shorter preparation time and high yields represent a definite
advantage over conventional stepwise solid-phase synthesis of vector–cargo molecules.
A plethora of chemoselective ligation methods is now available for the ligation
of unprotected peptides and proteins through amide and nonamide linkages; 116 many
of these may be applied to the production of translocating conjugates.
6.6 CATIONIC TRANSLOCATING PEPTIDES
GRKKRRQRRRPPQC
RQIKIWFQNRRMKWKK
RRWRRWWRRWWRRWRR
GWTLNPPGYLLGKINLKALAALAKKIL
PKKKRKV
KRPAATKKAGQAKKKL
AAVALLPAVLLALLAP
AAVLLPVLLAAP
Cationic translocating peptides are diverse and frequently used; the HIV Tat peptide
was one of the earliest translocation peptides described after it was determined that
the truncated peptide retained the translocation activity exhibited by the entire Tat
protein. 117 The amino acid sequence of the highly charged Tat peptide is shown in
Table 6.2. Able to localize rapidly to the nucleus once inside the cell, the Tat peptide
has also been shown to transport covalently bound peptides and proteins into cells
and tissues. 8 In one of the most notable studies, a Tat peptide/β-galactosidase fusion
protein was shown to translocate into cells derived from various murine tissue types.
The 120-kDa protein was also found to be enzymatically active within each different
cell type. 118 Other proteins successfully delivered to intracellular space by the Tat
peptide include p16INK, caspase3, RhoA, and p27kip1. 119-122
The penetratin peptide, also referred to as pAntp, is frequently used to deliver
peptides and small proteins into cells. This peptide is derived from a highly conserved
region of homeoprotein transcription factors. 123 The 16-residue penetratin is routinely
used as a vector (Table 6.2); however, internalization of a shorter 6-residue fragment
has also been reported. 124 Even though there is a lack of sequence homology between
penetratin and the Tat peptide, both consist predominantly of positively charged
residues. It seems that the translocation activity of penetratin is dependent upon the
distribution of the positively charged residues and two tryptophan residues that reside