Cystine Knot Peptides with Tuneable Activity and Mechanism

Li, Choi Yi; Rehm, Fabian B. H.; Yap, Kuok; Zdenek, Christina N.; Harding, Maxim D; Fry, Bryan G.; Durek, Thomas; Craik, David J.; Veer, Simon J. de

doi:10.1002/anie.202200951

Cited by 9 publications

(9 citation statements)

References 64 publications

(15 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Gurmarin contains three disulfide linkages, whereby a pair of disulfides form a loop through which the third disulfide bond passes, creating a heat-stable and protease-resistant structure known as an inhibitor cystine knot ( Craik et al., 2001 ). Knottin family of peptides is known to have diverse biological activities ( Wang et al., 2008 ; Parthasarathy et al., 2021 ; Li et al., 2022 ). Gurmarin peptide is structurally different from the autoinducing peptide (AIP) produced by Sa .…”

Section: Resultsmentioning

confidence: 99%

Inhibition of Staphylococcus aureus biofilm formation by gurmarin, a plant-derived cyclic peptide

Chang

Dowd

Brackee

et al. 2022

Front. Cell. Infect. Microbiol.

View full text Add to dashboard Cite

Staphylococcus aureus (Sa) is an opportunistic pathogen capable of causing various infections ranging from superficial skin infections to life-threatening severe diseases including pneumonia and sepsis. Sa produces biofilms readily on biotic and abiotic surfaces. Biofilm cells are embedded in a protective polysaccharide matrix and show an innate resistance to antibiotics, disinfectants, and clearance by host defenses. Additionally, biofilms serve as a source for systemic dissemination. Moreover, infections associated with biofilms may result in longer hospitalizations, a need for surgery, and may even result in death. Agents that inhibit the formation of biofilms and virulence without affecting bacterial growth to avoid the development of drug resistance could be useful for therapeutic purposes. In this regard, we identified and purified a small cyclic peptide, gurmarin, from a plant source that inhibited the formation of Sa biofilm under in vitro growth conditions without affecting the viability of the bacterium. The purified peptide showed a predicted molecular size of ~4.2 kDa on SDS-PAGE. Transcriptomic analysis of Sa biofilm treated with peptide showed 161 differentially affected genes at a 2-fold change, and some of them include upregulation of genes involved in oxidoreductases and downregulation of genes involved in transferases and hydrolases. To determine the inhibitory effect of the peptide against Sa biofilm formation and virulence in vivo, we used a rat-implant biofilm model. Sa infected implants with or without peptide were placed under the neck skin of rats for seven days. Implants treated with peptide showed a reduction of CFU and lack of edema and sepsis when compared to that of control animals without peptide. Taken together, gurmarin peptide blocks Sa biofilm formation in vitro and in vivo and can be further developed for therapeutic use.

show abstract

Section: Resultsmentioning

confidence: 99%

Inhibition of Staphylococcus aureus biofilm formation by gurmarin, a plant-derived cyclic peptide

Chang

Dowd

Brackee

et al. 2022

Front. Cell. Infect. Microbiol.

View full text Add to dashboard Cite

show abstract

“…Very recently, two MCoTI-based libraries were developed to guide the design of potent and selective serine protease inhibitors. A library of synthetic, acyclic knottins with sequence diversity in loop 1 identified that FXIIa prefers Phe derivatives at the P1′ position . As described earlier, FXIIa has also been screened by mRNA display using a genetically encoded MCoTI-II library that was produced by in vitro translation .…”

Section: Resultsmentioning

confidence: 99%

“…24 To improve the selectivity of [Q3R]M5, we examined substitutions in an adjacent loop (loop 1), which is regarded as the primary binding loop of MCoTI-II. We focused on two positions in the primed segment (P1′ and P2′) that were recently highlighted as important binding contacts 25,50 but are often overlooked in engineering studies. Substituting P2′ Leu with Gly maintained potent inhibition of FXIIa (K i = 2 nM) but was accompanied by a substantial loss in activity against matriptase, plasmin, and trypsin.…”

Section: ■ Discussionmentioning

confidence: 99%

“…A library of synthetic, acyclic knottins with sequence diversity in loop 1 identified that FXIIa prefers Phe derivatives at the P1′ position. 50 As described earlier, FXIIa has also been screened by mRNA display using a genetically encoded MCoTI-II library that was produced by in vitro translation. 25 This approach identified Gly as strongly preferred at the P2′ position.…”

Section: Mcoti-ii Variants With Fewer Residues In Loopmentioning

confidence: 99%

See 1 more Smart Citation

Engineering the Cyclization Loop of MCoTI-II Generates Targeted Cyclotides that Potently Inhibit Factor XIIa

et al. 2022

Self Cite

View full text Add to dashboard Cite

Factor XIIa (FXIIa) is a promising target for developing new drugs that prevent thrombosis without causing bleeding complications. A native cyclotide (MCoTI-II) is gaining interest for engineering FXIIa-targeted anticoagulants as this peptide inhibits FXIIa but not other coagulation proteases. Here, we engineered the native biosynthetic cyclization loop of MCoTI-II (loop 6) to generate improved FXIIa inhibitors. Decreasing the loop length led to gains in potency up to 7.7-fold, with the most potent variant having five residues in loop 6 (K i = 25 nM). We subsequently examined sequence changes within loop 6 and an adjacent loop, with substitutions at P4 and P2′ producing a potent FXIIa inhibitor (K i = 2 nM) that displayed more than 700-fold selectivity, was stable in human serum, and blocked the intrinsic coagulation pathway in human plasma. These findings demonstrate that engineering the biosynthetic cyclization loop can generate improved cyclotide variants, expanding their potential for drug discovery.

show abstract

“…An important class of DCPs, known as cystine knot peptides (CKP), are naturally found in various plants and animals, exhibiting diverse pharmacological activities [ 5 ]. CKPs typically consist of approximately 30–50 amino acids and contain six conserved cysteine residues, which form three disulfide bonds in a I-IV, II-V, III-VI arrangement.…”

Section: Introductionmentioning

confidence: 99%

Disulfide-constrained peptide scaffolds enable a robust peptide-therapeutic discovery platform

Zhou,

Cai,

et al. 2024

PLoS ONE

View full text Add to dashboard Cite

Peptides present an alternative modality to immunoglobulin domains or small molecules for developing therapeutics to either agonize or antagonize cellular pathways associated with diseases. However, peptides often suffer from poor chemical and physical stability, limiting their therapeutic potential. Disulfide-constrained peptides (DCP) are naturally occurring and possess numerous desirable properties, such as high stability, that qualify them as drug-like scaffolds for peptide therapeutics. DCPs contain loop regions protruding from the core of the molecule that are amenable to peptide engineering via direct evolution by use of phage display technology. In this study, we have established a robust platform for the discovery of peptide therapeutics using various DCPs as scaffolds. We created diverse libraries comprising seven different DCP scaffolds, resulting in an overall diversity of 2 x 1011. The effectiveness of this platform for functional hit discovery has been extensively evaluated, demonstrating a hit rate comparable to that of synthetic antibody libraries. By utilizing chemically synthesized and in vitro folded peptides derived from selections of phage displayed DCP libraries, we have successfully generated functional inhibitors targeting the HtrA1 protease. Through affinity maturation strategies, we have transformed initially weak binders against Notch2 with micromolar Kd values to high-affinity ligands in the nanomolar range. This process highlights a viable hit-to-lead progression. Overall, our platform holds significant potential to greatly enhance the discovery of peptide therapeutics.

show abstract

Cystine Knot Peptides with Tuneable Activity and Mechanism

Cited by 9 publications

References 64 publications

Inhibition of Staphylococcus aureus biofilm formation by gurmarin, a plant-derived cyclic peptide

Inhibition of Staphylococcus aureus biofilm formation by gurmarin, a plant-derived cyclic peptide

Engineering the Cyclization Loop of MCoTI-II Generates Targeted Cyclotides that Potently Inhibit Factor XIIa

Disulfide-constrained peptide scaffolds enable a robust peptide-therapeutic discovery platform

Contact Info

Product

Resources

About