Folding and aggregation of TEM β‐lactamase: Analogies with the formation of inclusion bodies in Escherichia coli

George Georgiou, Pascal Valax, Marc Ostermeier, Paul M. Horowitz

Resultado de la investigación: Articlerevisión exhaustiva

62 Citas (Scopus)


The enzyme TEM β‐lactamase has been used as a model for understanding the pathway leading to formation of inclusion bodies in Escherichia coli. The equilibrium denaturation of TEM β‐lactamase revealed that an intermediate that has lost enzymatic activity, native protein fluorescence, and UV absorption, but retains 60% of the native circular dichroism signal, becomes populated at intermediate (1.0–1.4 M) concentrations of guanidium chloride (GdmCl). This species exhibits a large increase in bis‐1‐anilino‐8‐naphthalene sulfonic acid fluorescence, indicating the presence of exposed hydrophobic surfaces. When TEM β‐lactamase was unfolded in different initial concentrations of GdmCl and refolded to the same final conditions by dialysis a distinct minimum in the yield of active protein was observed for initial concentrations of GdmCl in the 1.0–1.5 M range. It was shown that the lower reactivation yield was solely due to the formation of noncovalently linked aggregates. We propose that the aggregation of TEM β‐lactamase involves the association of a compact state having partially exposed hydrophobic surfaces. This hypothesis is consistent with our recent findings that TEM β‐lactamase inclusion bodies contains extensive secondary structure (Przybycien TM, Dunn JP, Valax P, Georgiou G, 1994, Protein Eng 7:131–136). Finally, we have also shown that protein aggregation was enhanced at higher temperatures and in the presence of 5 mM dithiothreitol and was inhibited by the addition of sucrose. These conditions exert a similar effect on the formation of inclusion bodies in vivo.

Idioma originalEnglish (US)
Páginas (desde-hasta)1953-1960
Número de páginas8
PublicaciónProtein Science
EstadoPublished - nov. 1994
Publicado de forma externa

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


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