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

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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

Fingerprint

Bacilli
Thermodynamic stability
Hot Temperature
Mutation
Thermolysin
Genetic Techniques
Membrane Proteins
Peptide Hydrolases
bacillolysin
Proteins

Keywords

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

ASJC Scopus subject areas

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

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.

The effect of engineering surface loops on the thermal stability of Bacillus subtilis neutral protease. / Hardy, F.; Vriend, G.; Van der Vinne, B.; Frigerio, F.; Grandi, G.; Venema, G.; Eijsink, V. G H.

In: Protein Engineering, Vol. 7, No. 3, 1994, p. 425-430.

Research output: Contribution to journalArticle

Hardy, F, Vriend, G, Van der Vinne, B, Frigerio, F, Grandi, G, Venema, G & Eijsink, VGH 1994, 'The effect of engineering surface loops on the thermal stability of Bacillus subtilis neutral protease', Protein Engineering, vol. 7, no. 3, pp. 425-430.
Hardy F, Vriend G, Van der Vinne B, Frigerio F, Grandi G, Venema G et al. The effect of engineering surface loops on the thermal stability of Bacillus subtilis neutral protease. Protein Engineering. 1994;7(3):425-430.
Hardy, F. ; Vriend, G. ; Van der Vinne, B. ; Frigerio, F. ; Grandi, G. ; Venema, G. ; Eijsink, V. G H. / The effect of engineering surface loops on the thermal stability of Bacillus subtilis neutral protease. In: Protein Engineering. 1994 ; Vol. 7, No. 3. pp. 425-430.
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