Structural requirements for the biosynthesis of backbone cyclic peptide libraries.
BACKGROUND
Combinatorial methods for the production of molecular libraries are an important source of ligand diversity for chemical biology. Synthetic methods focus on the production of small molecules that must traverse the cell membrane to elicit a response. Genetic methods enable intracellular ligand production, but products must typically be large molecules in order to withstand cellular catabolism. Here we describe an intein-based approach to biosynthesis of backbone cyclic peptide libraries that combines the strengths of synthetic and genetic methods.


RESULTS
Through site-directed mutagenesis we show that the DnaE intein from Synechocystis sp. PCC6803 is very promiscuous with respect to peptide substrate composition, and can generate cyclic products ranging from four to nine amino acids. Libraries with five variable amino acids and either one or four fixed residues were prepared, yielding between 10(7) and 10(8) transformants. The majority of randomly selected clones from each library gave cyclic products.


CONCLUSIONS
We have developed a versatile method for producing intracellular libraries of small, stable cyclic peptides. Genetic encoding enables facile manipulation of vast numbers of compounds, while low molecular weight ensures ready pharmacophore identification. The demonstrated flexibility of the method towards both peptide length and composition makes it a valuable addition to existing methods for generating ligand diversity.
Scott
C P
CP
Department of Chemistry, The Pennsylvania State University, University Park 16802, USA.
Abel-Santos
E
E
Jones
A D
AD
Benkovic
S J
SJ
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D003500:Cyclization; D004274:DNA, Recombinant / Q000235:genetics; D013058:Mass Spectrometry; D015394:Molecular Structure; D016296:Mutagenesis / Q000235:genetics; D019151:Peptide Library; D010456:Peptides, Cyclic / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D017384:Sequence Deletion / Q000235:genetics
D004274:DNA, Recombinant; D019151:Peptide Library; D010456:Peptides, Cyclic