title: Domains of Pyrococcus furiosus L-asparaginase fold sequentially and assemble through strong intersubunit associative forces.
abstract: Here, we report the folding and assembly of a Pyrococcus furiosus-derived protein, L-asparaginase (PfA). PfA functions as a homodimer, with each monomer made of distinct N- and C-terminal domains. The purified individual domains as well as single Trp mutant of each domain were subjected to chemical denaturation/renaturation and probed by combination of spectroscopic, chromatographic, quenching and scattering techniques. We found that the N-domain acts like a folding scaffold and assists the folding of remaining polypeptide. The domains displayed sequential folding with the N-domain having higher thermodynamic stability. We report that the extreme thermal stability of PfA is due to the presence of high intersubunit associative forces supported by extensive H-bonding and ionic interactions network. Our results proved that folding cooperativity in a thermophilic, multisubunit protein is dictated by concomitant folding and association of constituent domains directly into a native quaternary structure. This report gives an account of the factors responsible for folding and stability of a therapeutically and industrially important protein.
authors: Kusuma School of Biological Sciences, Indian Institute of Technology Delhi, Hauz Khas, 110016, New Delhi, India.
authors: Dushyant K
authors: DK
authors: Garg
authors: Rachana
authors: R
authors: Tomar
authors: Reema R
authors: RR
authors: Dhoke
authors: Ankit
authors: A
authors: Srivastava
authors: Bishwajit
authors: B
authors: Kundu
mesh_terms: D000595:Amino Acid Sequence; D001215:Asparaginase / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D004795:Enzyme Stability; D008969:Molecular Sequence Data; D017510:Protein Folding; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry; D020027:Pyrococcus furiosus / Q000201:enzymology*
chemical_list: D001426:Bacterial Proteins; D021122:Protein Subunits; D001215:Asparaginase