Artificial ribosomes for fully programmable synthesis of nonribosomal peptides

Abstract

Provided herein, in some embodiments, are artificial ribosomes that synthesize nonribosomal peptides, polyketides, and fatty acids with full control over peptide sequence. Also provided herein are methods for programmed synthesis of nonribosomal peptides, polyketides, and fatty acids. In particular, provided herein are methods for scalable synthesis of a wide range of antibacterial, antifungal, antiviral, and anticancer compounds.

1 .- 14 . (canceled) 
     
     
         15 . A molecular assembly line comprising:
 (a) a synthetic RNA scaffold that comprises at least two aptamers, wherein each aptamer comprises a polypeptide binding site; and   (b) at least two fusion proteins, each fusion protein comprising a polyketide synthase (PKS) or fatty acid synthase (FAS) module fused to a RNA-binding peptide that specifically binds to the polypeptide binding site of at least one aptamer within the synthetic RNA scaffold,   
       wherein the synthetic RNA scaffold and the at least two fusion proteins form a complex. 
     
     
         16 . The molecular assembly line of  claim 15 , wherein the synthetic RNA scaffold is a tRNA analog. 
     
     
         17 . The molecular assembly line of  claim 15 , wherein at least one fusion protein comprises a RNA-binding peptide selected from Lambda(G1N2R4), P22N, RevN7D, HTLV-1-Rex, and BIV-Tat. 
     
     
         18 . The molecular assembly line of  claim 17 , wherein at least one RNA binding peptide binds to an aptamer sequence within a tRNA analog. 
     
     
         19 . A synthetic nucleic acid sequence encoding the molecular assembly line of  claim 15 . 
     
     
         20 . A biological cell comprising the synthetic nucleic acid sequence of  claim 19 . 
     
     
         21 . A method for assembling the molecular assembly line of  claim 15 , the method comprising:
 (a) providing the molecular assembly line to a cell-free expression system; and   (b) incubating the cell-free system containing the molecular assembly line under conditions wherein the at least two fusion proteins are able to bind to the synthetic RNA scaffold.   
     
     
         22 . The method of  claim 21 , wherein the synthetic RNA scaffold comprises tRNA analogs. 
     
     
         23 . The method of  claim 21 , wherein at least one RNA-binding peptide comprises an RNA-binding domain from a protein selected from Lambda N, P22N, RevN7D, HTLV-1-Rex, and BIV-Tat. 
     
     
         24 . The method of  claim 23 , wherein at least one RNA-binding peptide binds to an aptamer sequence within a tRNA analog.