title: Discovery and characterization of pseudocyclic cystine-knot α-amylase inhibitors with high resistance to heat and proteolytic degradation.
abstract: Obesity and type 2 diabetes are chronic metabolic diseases, and those affected could benefit from the use of α-amylase inhibitors to manage starch intake. The pseudocyclics, wrightides Wr-AI1 to Wr-AI3, isolated from an Apocynaceae plant show promise for further development as orally active α-amylase inhibitors. These linear peptides retain the stability known for cystine-knot peptides in the presence of harsh treatment. They are resistant to heat treatment and endopeptidase and exopeptidase degradation, which is characteristic of cyclic cystine-knot peptides. Our NMR and crystallography analysis also showed that wrightides, which are currently the smallest proteinaceous α-amylase inhibitors reported, contain the backbone-twisting cis-proline, which is preceded by a nonaromatic residue rather than a conventional aromatic residue. The modeled structure and a molecular dynamics study of Wr-AI1 in complex with yellow mealworm α-amylase suggested that, despite having a similar structure and cystine-knot fold, the knottin-type α-amylase inhibitors may bind to insect α-amylase via a different set of interactions. Finally, we showed that the precursors of pseudocyclic cystine-knot α-amylase inhibitors and their biosynthesis in plants follow a secretory protein synthesis pathway. Together, our findings provide insights for the use of the pseudocyclic α-amylase inhibitors as useful leads for the development of orally active peptidyl bioactives, as well as an alternative scaffold for cyclic peptides for engineering metabolically stable human α-amylase inhibitors.
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore.
authors.forename: Phuong Q T
authors.initials: PQ
authors.lastname: Nguyen
authors.forename: Shujing
authors.initials: S
authors.lastname: Wang
authors.forename: Akshita
authors.initials: A
authors.lastname: Kumar
authors.forename: Li J
authors.initials: LJ
authors.lastname: Yap
authors.forename: Thuy T
authors.initials: TT
authors.lastname: Luu
authors.forename: Julien
authors.initials: J
authors.lastname: Lescar
authors.forename: James P
authors.initials: JP
authors.lastname: Tam
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D029062:Apocynaceae / Q000737:chemistry; D018360:Crystallography, X-Ray; D055808:Drug Discovery; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D006860:Hydrogen Bonding; D019476:Insect Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D011485:Protein Binding; D015202:Protein Engineering; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D012996:Solutions; D013712:Tenebrio / Q000201:enzymology; D000516:alpha-Amylases / Q000037:antagonists & inhibitors* / Q000737:chemistry
chemical_list: D004791:Enzyme Inhibitors; D019476:Insect Proteins; D010940:Plant Proteins; D012996:Solutions; D000516:alpha-Amylases
keywords: cis-proline; cystine knot; pseudocyclics; wrightide; α-amylase inhibitors