Soluble peptides having constrained, secondary conformation in solution and method of making same

A composition of matter comprising a plurality of E. coli cells containing a diverse population of expressible oligonucleotides, each of said oligonucleotides encoding a peptide having a desirable bias of predetermined covalent constraints, wherein each of said oligonucleotides is operationally linked to expression elements and to a nucleic acid encoding a coat protein contained within an E. coli filamentous bacteriophage vector so as to be expressed as a fusion protein or as a secreted peptide, said expressible oligonucleotides having a desirable bias of random codon sequences produced from random combinations of first and second oligonucleotide precursor populations, each or either of said first and second oligonucleotide precursor populations having a desirable bias of random codon sequences and wherein said plurality of cells comprise at least one of said expressible oligonucleotides which encodes a peptide with constrained secondary structure.

A kit for the preparation of vectors for the expression of a peptide:coat protein fusion protein or a secreted peptide from a diverse population of random peptides having a desirable bias of predetermined covalent constraints, wherein at least one of said random peptides in said diverse population is a peptide with constrained secondary structure, said peptides being generated from combined first and second oligonucleotide precursor populations which when combined have a desirable bias of random codon sequences, comprising:

• two E. coli filamentous bacteriophage vectors, a first filamentous bacteriophage vector having a cloning site for said first precursor oligonucleotides and a pair of restriction sites for operationally combining first precursor oligonucleotides with second precursor oligonucleotides;
• and a second filamentous bacteriophage vector having a cloning site for said second precursor oligonucleotides and a pair of restriction sites complementary to those on said first vector, one or both vectors containing expression elements capable of being operationally linked to said combined first and second precursor oligonucleotides and to a nucleic acid encoding a coat protein.

A cloning system for expressing as a peptide:coat protein fusion protein or as a secreted peptide a diverse population of oligonucleotides encoding random peptides having a desirable bias of predetermined covalent constraints, wherein at least one of said oligonucleotides within said diverse population encodes a peptide with constrained secondary structure, said oligonucleotides being generated from a desirable bias of random codon sequences, said cloning system comprising an E. coli filamentous bacteriophage vector having a pair of restriction sites so as to allow the operational combination of said oligonucleotides into a contiguous oligonucleotide encoding said peptide having constrained secondary structure, said contiguous oligonucleotide being operationally linked to expression elements and to a nucleic acid encoding a coat protein contained within an E. coli filamentous bacteriophage so as to be expressed as a peptide:coat protein fusion protein or as a secreted peptide.

A cloning system for expressing as a peptide:coat protein fusion protein or as a secreted peptide a diverse population of oligonucleotides encoding random peptides having a desirable bias of predetermined covalent constraints wherein at least one of said oligonucleotides within said diverse population encodes a peptide with constrained secondary structure, said oligonucleotides being generated from diverse populations of combined first and second oligonucleotide precursor populations each or either having a desirable bias of random codon sequences said cloning system, comprising:

• a set of first filamentous bacteriophage vectors having a diverse population of first precursor oligonucleotides having a desirable bias of random codon sequences and a second set of filamentous bacteriophage vectors having a diverse population of second precursor oligonucleotides having a desirable bias of random codon sequences, said first and second vectors each having a pair of restriction sites so as to allow the operational combination of said oligonucleotides into a contiguous oligonucleotide encoding said peptide having constrained secondary structure and to a nucleic acid encoding a bacteriophage coat protein.

A population of vectors comprising a diverse population of oligonucleotides operationally linked to (i) expression elements, (ii) a suppressible stop codon and (iii) a nucleic acid encoding a coat protein so as to be expressed as a fusion protein with said coat protein of an E. coli filamentous bacteriophage in a suppressor host or as a secreted peptide in a non-suppressor host, said oligonucleotides having a desirable bias of random codon sequences, and more than one codon encoding an amino acid capable of forming a covalent bond.