Biosynthesis and Biological Screening of a Genetically Encoded Library Based on the Cyclotide MCoTI-I

Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
ChemBioChem (Impact Factor: 3.06). 11/2009; 10(16):2663-70. DOI: 10.1002/cbic.200900534
Source: PubMed

ABSTRACT (Figure Presented) Cell-ing point: This study shows that MCoTI-cyclotides can provide an ideal scaffold for the biosynthesis of large combinatorial libraries inside living E. coli cells. Coupled to an appropriate in vivo reporter system, this library may rapidly be screened, for example, by fluorescence-activated cell sorting.

Download full-text


Available from: Julio A Camarero, Aug 29, 2015
  • Source
    • "These alterations may serve as a basis for the identification of sites that have target binding potential, i.e. sites 9 in Loop 1; 24, 26, 28 in Loop 5; and 30, 31, 33 in Loop 6. This prediction agrees with a study that showed that the aforementioned sites in Loops 1 and 5 play a role in binding with trypsin (Austin et al. 2009). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Cyclic proteins have evolved for millions of years across all kingdoms of life to confer structural stability over their acyclic counterparts while maintaining intrinsic functional properties. Here, we show that cyclic miniproteins (or peptides) from Momordica (Cucurbitaceae) seeds evolved in species that diverged from an African ancestor around 19 Ma. The ability to achieve head-to-tail cyclization of Momordica cyclic peptides appears to have been acquired through a series of mutations in their acyclic precursor coding sequences following recent and independent gene expansion event(s). Evolutionary analysis of Momordica cyclic peptides reveals sites that are under selection, highlighting residues that are presumably constrained for maintaining their function as potent trypsin inhibitors. Molecular dynamics of Momordica cyclic peptides in complex with trypsin reveals site-specific residues involved in target binding. In a broader context, this study provides a basis for selecting Momordica species to further investigate the biosynthesis of the cyclic peptides and for constructing libraries that may be screened against evolutionarily related serine proteases implicated in human diseases.
    Molecular Biology and Evolution 11/2014; 32(2). DOI:10.1093/molbev/msu307 · 14.31 Impact Factor
  • Source
    • "Cyclotides are also medium-sized polypeptides and therefore can be readily synthesized by standard solid-phase peptide-synthesis using either Boc-[54] or Fmoc-based [55] methodologies thus allowing the introduction of non-natural amino acids or other chemical modifications for lead optimization. They can also be encoded within standard cloning vectors and readily expressed in bacteria or animal cells [26] [70], thus making them ideal substrates for molecular evolution strategies to enable generation and selection of compounds with optimal binding and inhibitory characteristics using high throughput cellbased assays [106]. All of these characteristics make cyclotides appear as very promising leads or frameworks for development of peptide-based therapeutics and diagnostics [27] [68] [105] [107] Abbreviations "
    [Show abstract] [Hide abstract]
    ABSTRACT: Cyclotides are a growing family of large plant-derived backbone-cyclized polypeptides (≈30 amino acids long) that share a disulfide-stabilized core characterized by an unusual knotted structure. Their unique circular backbone topology and knotted arrangement of three disulfide bonds makes them exceptionally stable to thermal, chemical, and enzymatic degradation compared to other peptides of similar size. Currently more than 100 sequences of different cyclotides have been characterized and the number is expected to increase dramatically in the coming years. Considering their stability, biological activities and ability to cross the cell membrane, cyclotides can be exploited to develop new peptide-based drugs with high potential for success. The cyclotide scaffold can be engineered or evolved using molecular evolution to inhibit protein-protein interactions implicated in cancer and other human diseases, or design new antimicrobial. The present review reports the biological diversity and therapeutic potential of natural and engineered cyclotides.
    Current Molecular Pharmacology 11/2010; 3(3):153-63. DOI:10.2174/1874-470211003030153
  • [Show abstract] [Hide abstract]
    ABSTRACT: The formation of ultra-shallow junctions with low sheet resistance poses a major challenge in the scaling of advanced CMOS devices. Optimal device characteristics are achieved by a combination of low energy implants with low thermal budget activation. In this paper, the role of various thermal processes such as extension and source/drain anneal and deposition of nitrides and oxides on controlling junction parameters are examined. In addition, the effect of anneal ambient in the presence of an ultra-thin oxide layer is examined. Effect of junction profiles on device performance of a 150 nm CMOS is also reported. Improved transistor performance is obtained by minimizing thermal budget and optimizing the implanted extension dose
    Ion Implantation Technology, 2000. Conference on; 02/2000
Show more