Heat induces end to end repetitive association in P. furiosus L-asparaginase which enables its thermophilic property

Sci Rep. 2020 Dec 10;10(1):21702. doi: 10.1038/s41598-020-78877-z.

Abstract

It remains undeciphered how thermophilic enzymes display enhanced stability at elevated temperatures. Taking L-asparaginase from P. furiosus (PfA) as an example, we combined scattering shapes deduced from small-angle X-ray scattering (SAXS) data at increased temperatures with symmetry mates from crystallographic structures to find that heating caused end-to-end association. The small contact point of self-binding appeared to be enabled by a terminal short β-strand in N-terminal domain, Leu179-Val-Val-Asn182 (LVVN). Interestingly, deletion of this strand led to a defunct enzyme, whereas suplementation of the peptide LVVN to the defunct enzyme restored structural frameworkwith mesophile-type functionality. Crystal structure of the peptide-bound defunct enzyme showed that one peptide ispresent in the same coordinates as in original enzyme, explaining gain-of lost function. A second peptide was seen bound to the protein at a different location suggesting its possible role in substrate-free molecular-association. Overall, we show that the heating induced self-assembly of native shapes of PfA led to an apparent super-stable assembly.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Asparaginase / chemistry
  • Asparaginase / metabolism*
  • Crystallography, X-Ray
  • Enzyme Stability
  • Hot Temperature* / adverse effects
  • Protein Conformation, beta-Strand
  • Protein Denaturation
  • Protein Domains
  • Pyrococcus furiosus / enzymology*

Substances

  • Asparaginase