Luca Reggiani

Publications (2)8.2 Total impact

• Article: Proteolysis of the proofreading subunit controls the assembly of Escherichia coli DNA polymerase III catalytic core.

  Daniela Bressanin, Alessandra Stefan, Fabrizio Dal Piaz, Stefano Cianchetta, Luca Reggiani, Alejandro Hochkoeppler
  [show abstract] [hide abstract]
  ABSTRACT: The C-terminal region of the proofreading subunit (epsilon) of Escherichia coli DNA polymerase III is shown here to be labile and to contain the residues (identified between F187 and R213) responsible for association with the polymerase subunit (alpha). We also identify two alpha-helices of the polymerase subunit (comprising the residues E311-M335 and G339-D353, respectively) as the determinants of binding to epsilon. The C-terminal region of epsilon is degraded by the ClpP protease assisted by the GroL molecular chaperone, while other factors control the overall concentration in vivo of epsilon. Among these factors, the chaperone DnaK is of primary importance for preserving the integrity of epsilon. Remarkably, inactivation of DnaK confers to Escherichia coli inviable phenotype at 42 degrees C, and viability can be restored over-expressing epsilon. Altogether, our observations indicate that the association between epsilon and alpha subunits of DNA polymerase III depends on small portions of both proteins, the association of which is controlled by proteolysis of epsilon. Accordingly, the factors catalysing (ClpP, GroL) or preventing (DnaK) this proteolysis exert a crucial checkpoint of the assembly of Escherichia coli DNA polymerase III core.
  Biochimica et Biophysica Acta 08/2009; 1794(11):1606-15. · 4.66 Impact Factor
• Article: Silencing of the gene coding for the epsilon subunit of DNA polymerase III slows down the growth rate of Escherichia coli populations.

  Alessandra Stefan, Luca Reggiani, Stefano Cianchetta, Annalisa Radeghieri, Antonio Gonzalez Vara y Rodriguez, Alejandro Hochkoeppler
  [show abstract] [hide abstract]
  ABSTRACT: Chromosome replication in Escherichia coli is accomplished by the multimeric enzyme DNA polymerase III; the relevance, in vivo, of the epsilon subunit (encoded by dnaQ) for processivity and fidelity of DNA polymerase III has been evaluated. To this aim, dnaQ has been conditionally silenced by means of in vivo expression of different antisense RNAs. Unexpectedly, the presence of the Shine-Dalgarno sequence is essential for the effectiveness of antisense constructs. Silencing of dnaQ induces a severe decrease in growth rate not paralleled by high mutation frequencies, suggesting that the epsilon subunit primarily affects the processivity of DNA polymerase III.
  FEBS Letters 08/2003; 546(2-3):295-9. · 3.54 Impact Factor