Your browser doesn't support javascript.
loading
Crystal structure and physicochemical characterization of a phytocystatin from Humulus lupulus: Insights into its domain-swapped dimer.
Moura, Gustavo Trajano de; Souza, Amanda Araújo; Garay, Aisel Valle; Freitas, Sonia Maria de; Valadares, Napoleão Fonseca.
Affiliation
  • Moura GT; Laboratório de Biofísica Molecular, Departamento de Biologia Celular, Universidade de Brasília, Brasília, 70910-900, Brazil.
  • Souza AA; Laboratório de Biofísica Molecular, Departamento de Biologia Celular, Universidade de Brasília, Brasília, 70910-900, Brazil.
  • Garay AV; Laboratório de Biofísica Molecular, Departamento de Biologia Celular, Universidade de Brasília, Brasília, 70910-900, Brazil.
  • Freitas SM; Laboratório de Biofísica Molecular, Departamento de Biologia Celular, Universidade de Brasília, Brasília, 70910-900, Brazil.
  • Valadares NF; Laboratório de Biofísica Molecular, Departamento de Biologia Celular, Universidade de Brasília, Brasília, 70910-900, Brazil. Electronic address: napo@unb.br.
Biochim Biophys Acta Proteins Proteom ; 1869(1): 140541, 2021 01.
Article in En | MEDLINE | ID: mdl-32947025
ABSTRACT
Phytocystatins are a family of plant cysteine-protease inhibitors of great interest due to their biotechnological application in culture improvement. It was shown that their expression in plants increases resistance to herbivory by insects and improves tolerance to both biotic and abiotic stress factors. In this work, owing to the economical relevance of the source organism, a phytocystatin from hop (Humulus lupulus), Hop1, was produced by heterologous expression in E. coli Lemo21 (DE3) cultivated in auto-inducing ZYM-5052 medium and purified by immobilized metal ion affinity and size exclusion chromatography. Thermal denaturation assays by circular dichroism showed that Hop1 exhibited high melting temperatures ranging from 82 °C to 85 °C and high thermal stability at a wide pH range, with ΔG25's higher than 12 kcal/mol. At 20 °C and pH 7.6, the dimeric conformation of the protein is favored according to size exclusion chromatography and analytical ultracentrifugation data, although monomers and higher order oligomers could still be detected in a lesser extent. The crystal structure of Hop1 was solved in the space groups P 2 21 21 and C 2 2 21 at resolutions of 1.80 Å and 1.68 Å, respectively. In both models, Hop1 is folded as a domain-swapped dimer where the first inhibitory loop undergoes a significant structural change and interacts with their equivalent from the other monomer forming a long antiparallel beta strand, leading to loss of inhibitory activity.
Subject(s)
Key words

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Plant Proteins / Cystatins / Cysteine Proteinase Inhibitors / Humulus Type of study: Prognostic_studies Language: En Journal: Biochim Biophys Acta Proteins Proteom Year: 2021 Document type: Article Affiliation country: Brazil

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Plant Proteins / Cystatins / Cysteine Proteinase Inhibitors / Humulus Type of study: Prognostic_studies Language: En Journal: Biochim Biophys Acta Proteins Proteom Year: 2021 Document type: Article Affiliation country: Brazil