The effect of engineering surface loops on the thermal stability of Bacillus subtilis neutral protease

F. Hardy, G. Vriend, B. Van der Vinne, F. Frigerio, G. Grandi, G. Venema, V. G H Eijsink

Research output: Contribution to journalArticle

Abstract

Using genetic techniques the contribution of surface loops to the thermal stability of Bacillus subtilis neutral protease (NP-sub) was studied. Mutations were designed to make the surface of NP-sub more similar to the surface of more thermostable neutral proteases such as thermolysin (TLN). The mutations included the replacement of an irregular loop by a shorter variant and the introduction of a ten-residue β-hairpin. In general, these drastic mutations had little effect on the production and activity of NP-sub, indicating the feasibility of major structural rearrangements at the surface of proteins. In the most stable mutant, exhibiting an increase in thermal stability of 1.1°C, ~10% of the surface of NP-sub was modified. Several NP-sub variants carrying multiple mutations were constructed. Non-additive effects on thermal stability were observed, which were interpreted on the basis of a model for thermal inactivation, that emphasizes the importance of local unfolding processes for thermal stability.

Original languageEnglish
Pages (from-to)425-430
Number of pages6
JournalProtein Engineering
Volume7
Issue number3
Publication statusPublished - 1994

Keywords

  • Bacillus
  • Loop
  • Neutral protease
  • Surface
  • Thermal stability

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Pharmacology
  • Neuroscience(all)
  • Immunology and Microbiology(all)
  • Bioengineering
  • Biotechnology

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  • Cite this

    Hardy, F., Vriend, G., Van der Vinne, B., Frigerio, F., Grandi, G., Venema, G., & Eijsink, V. G. H. (1994). The effect of engineering surface loops on the thermal stability of Bacillus subtilis neutral protease. Protein Engineering, 7(3), 425-430.