Biosynthesis, Regulation and Export of Lasso Peptides - Lasso Peptides: Bacterial Strategies to Make and Maintain Bioactive Entangled Scaffolds - page 79

Chapter 4
Biosynthesis, Regulation and Export
of Lasso Peptides

4.1 Biosynthesis of Lasso Peptides

Lasso peptides composed of 15-24 amino acids share an interlocked topology,
which consists of an N-terminal macrolactam ring threaded by a C-terminal tail that
remains locked inside. This molecular architecture remains a challenge for chem-
ists. Indeed, no group in the world has succeeded in the chemical synthesis of such a
constrained and entropically disfavoured topology. However, bacteria can establish
this fascinating structure, thanks to specific enzymes that are capable of transform-
ing a linear precursor made of unmodified amino acids into the lasso topology. The
first in vitro reconstitution of microcin J25 (MccJ25) biosynthesis (Duquesne et al.
2007 ) opened the way to study the molecular mechanism in detail. It was demon-
strated that the lasso topology was formed upon posttranslational modification of a
58-amino acid precursor (McjA) by two enzymes McjB and McjC encoded in the
MccJ25 gene cluster, in the presence of adenosine triphosphate (ATP) and Mg 2+
ions (Duquesne et al. 2007 ) (Fig. 4.1 ).

4.1.1 Maturation Enzymes

The biosynthesis of lasso peptides follows the same logic as other ribosomally
synthesized and posttranslationally modified peptides (RiPPs; Arnison et al. 2013 ;
Yang and van der Donk 2013 ), i.e. they are synthesized as linear precursor peptides,
which are further subjected to posttranslational modifications and transformed into
the mature form. Being the only member with a characterized genetic system until
2008, MccJ25 served for many years and still serves as a model for studying lasso
peptide biosynthesis. The biosynthetic machinery is composed of three proteins
McjA, McjB and McjC. The precursor peptide McjA contains a 37-amino acid (aa)
leader peptide fused at the N-terminus of the core sequence. Two chemical modi-
fications on McjA, comprising both leader peptide cleavage and macrocyclization,
give rise to mature MccJ25 with a defined lasso topology. In vitro experiments
using recombinant proteins (Duquesne et al. 2007 ) or extracts from cells express-