Full Sequence Amino Acid Scanning of θ‑Defensin RTD‑1 Yields a Potent Anthrax Lethal Factor Protease Inhibitor

θ-Defensin RTD-1 is a noncompetitive
inhibitor of anthrax
lethal factor (LF) protease (IC50 = 390 ± 20 nM, Ki = 365 ± 20 nM) and a weak inhibitor of
other mammalian metalloproteases such as TNFα converting enzyme
(TACE) (Ki = 4.45 ± 0.48 μM).
Using full sequence amino acid scanning in combination with a highly
efficient “one-pot” cyclization-folding approach, we
obtained an RTD-1-based peptide that was around 10 times more active
than wild-type RTD-1 in inhibiting LF protease (IC50 =
43 ± 3 nM, Ki = 18 ± 1 nM).
The most active peptide was completely symmetrical, rich in Arg and
Trp residues, and able to adopt a native RTD-1-like structure. These
results show the power of optimized chemical peptide synthesis approaches
for the efficient production of libraries of disulfide-rich backbone-cyclized
peptides to quickly perform structure–activity relationship
studies for optimizing protease inhibitors.