Reduced stability and enhanced surface hydrophobicity drive the binding of apo-aconitase with GroEL during chaperone assisted refolding |
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| Authors: | Parul Gupta Saroj Mishra Tapan Kumar Chaudhuri |
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| Affiliation: | 1. Departamento de Biotecnología y Bioingeniería, Centro de Investigación y de Estudios Avanzados del IPN, Col. San Pedro Zacatenco, Mexico, DF 07360, Mexico;2. Laboratorio de Investigación Bioquímica, ENMyH-IPN, Guillermo Massieu Helguera No. 239, La Escalera Ticoman, Mexico,DF 07320, Mexico;3. Departamento de Infectómica y Patogénesis Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Col. San Pedro Zacatenco, Mexico, DF 07360, Mexico;1. Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126 Milan, Italy;2. Consorzio Nazionale Interuniversitario per le Scienze Fisiche della Materia (CNISM), UdR Milano-Bicocca, Milan, Italy;3. Department of Plant Molecular Biology, Faculty of Biology and Medicine, University of Lausanne, 1015 Lausanne, Switzerland;4. Department of Physics, University of Milano-Bicocca, Piazza della Scienza 3, 20126 Milan, Italy;1. Unidad de Biofisica (CSIC-UPV/EHU) and Departmento de Bioquímica y Biología Molecular, Facultad de Ciencia y Tecnología, Universidad del País Vasco (UPV/EHU), Aptdo. 644, 48080 Bilbao, Spain;2. Centro de Investigaciones Biológicas, CSIC, C/ Ramiro de Maeztu 9, E-40 28040 Madrid, Spain;1. School of Biotechnology, Jawaharlal Nehru University, New Delhi 110067, India;2. Bioinformatics Section, Department of Computer Sciences, Hamdard University, New Delhi 110062;1. School of Chemistry, University of Hyderabad, Hyderabad 500046, India;2. Animal Reproduction Laboratory, ICAR-National Research Centre on Equines, Bikaner 334001, India |
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| Abstract: | Apo-aconitase, the Fe4S4 cluster free form of TCA cycle enzyme aconitase, binds with GroEL and dissociates itself upon maturation through insertion of the cluster. It is not clearly established as to why apo-protein binds with GroEL. In order to explore the possibility that stability is a factor responsible for the aggregation of apo-form at low ionic strengths and hence it associates with GroEL to avoid the unfavorable event, we carried out the unfolding studies with holo- and apo-aconitase. By probing the unfolding process through the changes in secondary structural element, exposed surface hydrophobicity, and the microenvironment around tryptophan residues, we were able to establish the relevant changes associated with the event. Apparent guanidine hydrochloride concentration required for unfolding of 50% of aconitase indicates that aconitase is destabilized in the absence of the Fe4S4 cluster. The destabilization of the apo-aconitase was further reflected through its three times higher rate of unfolding as compared to the holo-protein. It was also observed that the apo-form has higher surface hydrophobicity than the holo-form. Hence, the lower ground state stability and higher solvent exposed hydrophobic surface of the apo-form makes it aggregation prone. Based on the present observation and earlier findings, we propose that binding of apo-aconitase to GroEL not only rescues it from the aggregation, but also assists in the final stage of maturation by orienting the cluster insertion site of GroEL bound apo-protein. This information sheds new light on the potential role of GroEL in the biosynthetic pathway of the metallo proteins. |
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