title: The TCP1γ subunit of Leishmania donovani forms a biologically active homo-oligomeric complex.
abstract: Chaperonins are a class of molecular chaperons that encapsulate nascent or stress-denatured proteins and assist their intracellular assembly and folding in an ATP-dependent manner. The ubiquitous eukaryotic chaperonin, TCP1 ring complex is a hetero-oligomeric complex comprising two rings, each formed of eight subunits that may have distinct substrate recognition and ATP hydrolysis properties. In Leishmania, only the TCP1γ subunit has been cloned and characterized. It exhibited differential expression at various growth stages of promastigotes. In the present study, we expressed the TCP1γ subunit in Escherichia coli to investigate whether it forms chaperonin-like complexes and plays a role in protein folding. LdTCP1γ formed high-molecular-weight complexes within E. coli cells as well as in Leishmania cell lysates. The recombinant protein is arranged into two back-to-back rings of seven subunits each, as predicted by homology modelling and observed by negative staining electron microscopy. This morphology is consistent with that of the oligomeric double-ring group I chaperonins found in mitochondria. The LdTCP1γ homo-oligomeric complex hydrolysed ATP, and was active as assayed by luciferase refolding. Thus, the homo-oligomer performs chaperonin reactions without partner subunit(s). Further, co-immunoprecipitation studies revealed that LdTCP1γ interacts with actin and tubulin proteins, suggesting that the complex may have a role in maintaining the structural dynamics of the cytoskeleton of parasites.
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute, Lucknow, India.|Academy of Scientific and Innovative Research, Anusandhan Bhawan, New Delhi, India.
authors.lastname: Bhaskar
authors.affiliation: Academy of Scientific and Innovative Research, Anusandhan Bhawan, New Delhi, India.|Electron Microscopy Unit, Sophisticated Analytical Instrument Facility, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Kalyan
authors.initials: K
authors.lastname: Mitra
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Jitendra
authors.initials: J
authors.lastname: Kuldeep
authors.affiliation: Academy of Scientific and Innovative Research, Anusandhan Bhawan, New Delhi, India.|Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, Lucknow, India.
authors.forename: Mohammad Imran
authors.initials: MI
authors.lastname: Siddiqi
authors.affiliation: Biochemistry Division, CSIR-Central Drug Research Institute, Lucknow, India.|Academy of Scientific and Innovative Research, Anusandhan Bhawan, New Delhi, India.
authors.forename: Neena
authors.initials: N
authors.lastname: Goyal
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D056404:Chaperonin Containing TCP-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000166:cytology / Q000378:metabolism; D007893:Leishmania donovani / Q000737:chemistry* / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*
chemical_list: D000199:Actins; D021122:Protein Subunits; D014404:Tubulin; D056404:Chaperonin Containing TCP-1
keywords: Leishmania donovani; T‐complex protein 1 γ subunit; chaperonin; co‐immunoprecipitation; in vitro protein folding