Identification of the substrate specificity-conferring amino acid residues of 4-coumarate:coenzyme A ligase allows the rational design of mutant enzymes with new catalytic properties

The Journal of Biological Chemistry
Hans-Peter Stuible, Erich Kombrink

Abstract

4-Coumarate:coenzyme A ligases (4CLs) generally use, in addition to coumarate, caffeate and ferulate as their main substrates. However, the recently cloned Arabidopsis thaliana isoform At4CL2 is exceptional because it has no appreciable activity with ferulate. On the basis of information obtained from the crystal structure of the phenylalanine-activating domain of gramicidin S-synthetase, 10 amino acid residues were identified that may form the substrate binding pocket of 4CL. Among these amino acids, representing the putative "substrate specificity motif," only one residue, Met(293), was not conserved in At4CL2, compared with At4CL1 and At4CL3, two isoforms using ferulate. Substitution of Met(293) or Lys(320), another residue of the putative substrate specificity motif, which in the predicted three-dimensional structure is located in close proximity to Met(293), by smaller amino acids converted At4CL2 to an enzyme capable of using ferulate. The activity with caffeate was not or only moderately affected. Conversely, substitution of Met(293) by bulky aromatic amino acids increased the apparent affinity (K(m)) for caffeate up to 10-fold, whereas single substitutions of Val(294) did not affect substrate use. The results support ou...Continue Reading

References

May 1, 1975·Zeitschrift Für Naturforschung. Section C: Biosciences·J Stöckigt, M H Zenk
Mar 17, 1975·European Journal of Biochemistry·K H Knobloch, K Hahlbrock
Jan 11, 1984·Nucleic Acids Research·J DevereuxO Smithies
Mar 17, 1995·The Journal of Biological Chemistry·T Stachelhaus, M A Marahiel
Jun 6, 1998·Proceedings of the National Academy of Sciences of the United States of America·W J HuV L Chiang
Mar 6, 1999·Current Opinion in Plant Biology·B Weisshaar, G I Jenkins
Jul 27, 1999·The Plant Journal : for Cell and Molecular Biology·J EhltingE Kombrink
Jul 28, 1999·Chemistry & Biology·T StachelhausM A Marahiel
Sep 28, 1999·Trends in Plant Science·R A Dixon, C L Steele
Oct 6, 1999·Chemistry & Biology·H von DöhrenM Pavela-Vrancic

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Citations

Feb 4, 2012·The Arabidopsis Book·Christopher M Fraser, Clint Chapple
Sep 29, 2001·The Plant Journal : for Cell and Molecular Biology·J EhltingC J Douglas
Feb 22, 2002·European Journal of Biochemistry·Christian LindermayrJürgen Ebel
May 17, 2013·Plant & Cell Physiology·Shubhra RastogiAjit K Shasany
Mar 20, 2003·Plant Physiology·Małgorzata Pietrowska-BorekAndrzej Guranowski
Mar 12, 2011·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Hiroyuki MoritaIkuro Abe
Dec 18, 2002·Journal of Bacteriology·Masafumi KanekoSueharu Horinouchi
Jan 11, 2014·Functional & Integrative Genomics·Kalenahalli N YogendraTeresa Mosquera
Sep 11, 2002·Proceedings of the National Academy of Sciences of the United States of America·Jurgen J MayMilton T Stubbs
Jun 24, 2003·Proceedings of the National Academy of Sciences of the United States of America·Katja SchneiderHans-Peter Stuible
Feb 11, 2004·Proceedings of the National Academy of Sciences of the United States of America·Björn Hamberger, Klaus Hahlbrock
Apr 18, 2014·Biological & Pharmaceutical Bulletin·Yeon Bok KimSang Un Park
Feb 19, 2011·Journal of Plant Physiology·Mariam M GaidLudger Beerhues
Sep 3, 2011·The Plant Journal : for Cell and Molecular Biology·Moti AlbersteinHagai Abeliovich
Apr 14, 2016·Biochimica Et Biophysica Acta·Rémi DulermoJean-Marc Nicaud
Apr 23, 2016·Photochemical & Photobiological Sciences : Official Journal of the European Photochemistry Association and the European Society for Photobiology·R A PradoV R Viviani
May 31, 2017·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Sang-Hoon LeeYeon Bok Kim
Jul 3, 2013·International Journal of Molecular Sciences·Teresa DocimoMonica Mattana
Jan 10, 2018·Frontiers in Plant Science·Hye Lin ParkMan-Ho Cho
Oct 6, 2017·Journal of Experimental Botany·Hongjie FengJie Sun
Jan 7, 2021·Functional & Integrative Genomics·Douglas Jardim-MessederGilberto Sachetto-Martins
Jun 1, 2021·Chemical & Pharmaceutical Bulletin·Takahiro MoriIkuro Abe

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