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A multidomain PARP14 construct suitable for bacterial expression

Constantinos Chatzicharalampous1, Herwig Schüler1

  • 1Division of Biochemistry and Structural Biology, Department of Chemistry, Lund University, SE-22362, Lund, Sweden.

Protein Expression and Purification
|August 18, 2024
PubMed

Insights

Researchers developed a new method to produce functional Poly-ADP-ribose polymerase-14 (PARP14) in bacteria. This breakthrough enables crucial structural and functional studies of PARP14, a protein linked to cancer and viral infections.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Enzymology

Background:

  • Poly-ADP-ribose polymerase-14 (PARP14) is implicated in cancer and inflammation.
  • PARP14's diverse domains are involved in viral infection processes.
  • Previous studies faced challenges in obtaining sufficient pure PARP14 for in vitro analysis.

Purpose of the Study:

  • To develop a strategy for bacterial expression and purification of a functional PARP14 multidomain construct.
  • To create a PARP14 protein amenable to structural and functional investigations.

Main Methods:

  • Engineered a PARP14 construct by replacing an internal KH domain and unstructured region with a SUMO domain.
  • Utilized bacterial expression systems for protein production.
  • Purified the modified multidomain PARP14 construct.

Main Results:

  • Successfully expressed and purified a functional multidomain PARP14 construct in bacteria.
  • The engineered construct retained essential ADP-ribosyltransferase and de-MARylase activities.
  • The purified construct is suitable for further structural and functional studies.

Conclusions:

  • A novel bacterial expression and purification strategy for PARP14 has been established.
  • The developed PARP14 construct maintains key enzymatic activities.
  • This work facilitates advanced research into PARP14's role in disease and biological processes.
Keywords:
ADP-Ribose bindingADP-Ribosyl glycohydrolaseADP-RibosyltransferaseBacterial expressionMono-ADP-RibosylationPoly-ADP-Ribose polymerase

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