Conformational mobility of active and E-64-inhibited actinidin
Abstract
Background: Actinidin, a protease from kiwifruit, belongs to the C1 family of cysteine proteases. Cysteine proteases were found to be involved in many disease states and are valid therapeutic targets. Actinidin has a wide pH activity range and wide substrate specificity, which makes it a good model system for studying enzyme-substrate interactions. Methods: The influence of inhibitor (E-64) binding on the conformation of actinidin was examined by 2D PAGE, circular dichroism (CD) spectroscopy, hydrophobic ligand binding assay, and molecular dynamics simulations. Results: Significant differences were observed in electrophoretic mobility of proteolytically active and E-64-inhibited actinidin. CD spectrometry and hydrophobic ligand binding assay revealed a difference in conformation between active and inhibited actinidin. Molecular dynamics simulations showed that a loop defined by amino-acid residues 88-104 had greater conformational mobility in the inhibited enzyme than in the active one.... During MD simulations, the covalently bound inhibitor was found to change its conformation from extended to folded, with the guanidino moiety approaching the carboxylate. Conclusions: Conformational mobility of actinidin changes upon binding of the inhibitor, leading to a sequence of events that enables water and ions to protrude into a newly formed cavity of the inhibited enzyme. Drastic conformational mobility of E-64, a common inhibitor of cysteine proteases found in many crystal structures stored in PDB, was also observed. General significance: The analysis of structural changes which occur upon binding of an inhibitor to a cysteine protease provides a valuable starting point for the future design of therapeutic agents.
Keywords:
Actinidin / E-64 / Molecular dynamics / Cysteine proteaseSource:
Biochimica et Biophysica Acta: General Subjects, 2013, 1830, 10, 4790-4799Publisher:
- Elsevier Science Bv, Amsterdam
Funding / projects:
- Allergens, antibodies, enzymes and small physiologically important molecules: design, structure, function and relevance (RS-172049)
- Rational design and synthesis of biologically active and coordination compounds and functional materials, relevant for (bio)nanotechnology (RS-172035)
- Reinforcement of the Faculty of Chemistry, University of Belgrade, towards becoming a Center of Excellence in the region of WB for Molecular Biotechnology and Food research (EU-256716)
- High-Performance Computing Infrastructure for South East Europe's Research Communities (EU-261499)
Note:
- Supplementary material: http://cherry.chem.bg.ac.rs/handle/123456789/3476
DOI: 10.1016/j.bbagen.2013.06.015
ISSN: 0304-4165
PubMed: 23803410
WoS: 000323854900041
Scopus: 2-s2.0-84880183998
Collections
Institution/Community
Hemijski fakultet / Faculty of ChemistryTY - JOUR AU - Grozdanović, Milica M. AU - Drakulić, Branko J. AU - Gavrović-Jankulović, Marija PY - 2013 UR - https://cherry.chem.bg.ac.rs/handle/123456789/1398 AB - Background: Actinidin, a protease from kiwifruit, belongs to the C1 family of cysteine proteases. Cysteine proteases were found to be involved in many disease states and are valid therapeutic targets. Actinidin has a wide pH activity range and wide substrate specificity, which makes it a good model system for studying enzyme-substrate interactions. Methods: The influence of inhibitor (E-64) binding on the conformation of actinidin was examined by 2D PAGE, circular dichroism (CD) spectroscopy, hydrophobic ligand binding assay, and molecular dynamics simulations. Results: Significant differences were observed in electrophoretic mobility of proteolytically active and E-64-inhibited actinidin. CD spectrometry and hydrophobic ligand binding assay revealed a difference in conformation between active and inhibited actinidin. Molecular dynamics simulations showed that a loop defined by amino-acid residues 88-104 had greater conformational mobility in the inhibited enzyme than in the active one. During MD simulations, the covalently bound inhibitor was found to change its conformation from extended to folded, with the guanidino moiety approaching the carboxylate. Conclusions: Conformational mobility of actinidin changes upon binding of the inhibitor, leading to a sequence of events that enables water and ions to protrude into a newly formed cavity of the inhibited enzyme. Drastic conformational mobility of E-64, a common inhibitor of cysteine proteases found in many crystal structures stored in PDB, was also observed. General significance: The analysis of structural changes which occur upon binding of an inhibitor to a cysteine protease provides a valuable starting point for the future design of therapeutic agents. PB - Elsevier Science Bv, Amsterdam T2 - Biochimica et Biophysica Acta: General Subjects T1 - Conformational mobility of active and E-64-inhibited actinidin VL - 1830 IS - 10 SP - 4790 EP - 4799 DO - 10.1016/j.bbagen.2013.06.015 ER -
@article{ author = "Grozdanović, Milica M. and Drakulić, Branko J. and Gavrović-Jankulović, Marija", year = "2013", abstract = "Background: Actinidin, a protease from kiwifruit, belongs to the C1 family of cysteine proteases. Cysteine proteases were found to be involved in many disease states and are valid therapeutic targets. Actinidin has a wide pH activity range and wide substrate specificity, which makes it a good model system for studying enzyme-substrate interactions. Methods: The influence of inhibitor (E-64) binding on the conformation of actinidin was examined by 2D PAGE, circular dichroism (CD) spectroscopy, hydrophobic ligand binding assay, and molecular dynamics simulations. Results: Significant differences were observed in electrophoretic mobility of proteolytically active and E-64-inhibited actinidin. CD spectrometry and hydrophobic ligand binding assay revealed a difference in conformation between active and inhibited actinidin. Molecular dynamics simulations showed that a loop defined by amino-acid residues 88-104 had greater conformational mobility in the inhibited enzyme than in the active one. During MD simulations, the covalently bound inhibitor was found to change its conformation from extended to folded, with the guanidino moiety approaching the carboxylate. Conclusions: Conformational mobility of actinidin changes upon binding of the inhibitor, leading to a sequence of events that enables water and ions to protrude into a newly formed cavity of the inhibited enzyme. Drastic conformational mobility of E-64, a common inhibitor of cysteine proteases found in many crystal structures stored in PDB, was also observed. General significance: The analysis of structural changes which occur upon binding of an inhibitor to a cysteine protease provides a valuable starting point for the future design of therapeutic agents.", publisher = "Elsevier Science Bv, Amsterdam", journal = "Biochimica et Biophysica Acta: General Subjects", title = "Conformational mobility of active and E-64-inhibited actinidin", volume = "1830", number = "10", pages = "4790-4799", doi = "10.1016/j.bbagen.2013.06.015" }
Grozdanović, M. M., Drakulić, B. J.,& Gavrović-Jankulović, M.. (2013). Conformational mobility of active and E-64-inhibited actinidin. in Biochimica et Biophysica Acta: General Subjects Elsevier Science Bv, Amsterdam., 1830(10), 4790-4799. https://doi.org/10.1016/j.bbagen.2013.06.015
Grozdanović MM, Drakulić BJ, Gavrović-Jankulović M. Conformational mobility of active and E-64-inhibited actinidin. in Biochimica et Biophysica Acta: General Subjects. 2013;1830(10):4790-4799. doi:10.1016/j.bbagen.2013.06.015 .
Grozdanović, Milica M., Drakulić, Branko J., Gavrović-Jankulović, Marija, "Conformational mobility of active and E-64-inhibited actinidin" in Biochimica et Biophysica Acta: General Subjects, 1830, no. 10 (2013):4790-4799, https://doi.org/10.1016/j.bbagen.2013.06.015 . .