Related Experiment Video
Updated: Jul 30, 2026
08:59
Defining Substrate Specificities for Lipase and Phospholipase Candidates
Published on: November 23, 2016
Creating lipoxygenases with new positional specificities by site-directed mutagenesis
1Institute of Plant Genetics and Crop Plant Research, D-06466 Gatersleben, Germany.
Biochemical Society Transactions
|February 15, 2001
Summary
Researchers identified key amino acid residues that control plant lipoxygenase (LOX) positional specificity. Altering a single residue generated novel LOX enzymes with changed specificity, advancing our understanding of plant LOX function.
Area of Science:
- Biochemistry
- Enzymology
- Plant Molecular Biology
Background:
- Plant lipoxygenases (LOXs) are enzymes involved in various plant processes.
- Understanding the determinants of LOX positional specificity is crucial for elucidating their functions.
- Previous studies primarily focused on mammalian LOXs, identifying two regions with specificity determinants.
Purpose of the Study:
- To identify amino acid residues responsible for altering the positional specificity of plant lipoxygenases (LOXs).
- To generate novel plant LOX variants with modified positional specificities through targeted amino acid alterations.
Main Methods:
- Performed multiple sequence alignments of plant LOX enzymes.
- Utilized structural modeling to analyze enzyme-substrate interactions.
- Employed site-directed mutagenesis to alter specific amino acid residues within the LOX active site.
Main Results:
- Identified three primary amino acid residues as key determinants of plant LOX positional specificity.
- Successfully generated two novel plant LOX variants: a gamma-linolenate 6-LOX and an arachidonate 11-LOX.
- Demonstrated that altering a single amino acid residue within the active site can significantly change LOX positional specificity.
Conclusions:
- A single amino acid change in the active site is sufficient to confer new positional specificities to plant LOXs.
- The identified amino acid determinants provide valuable insights into the mechanisms governing plant LOX substrate recognition.
- This study offers a foundation for engineering plant LOXs with tailored specificities for research and biotechnological applications.
Related Concept Videos
02:40Ligand Binding Sites
Proteins are dynamic macromolecules that carry out a wide variety of essential processes; however, the activities of most proteins depend on their interactions with other molecules or ions, known as ligands.
Protein-ligand interactions are quite specific; even though numerous potential ligands surround a cellular protein at any given time, only a particular ligand can bind to that protein. Moreover, a ligand binds only to a dedicated area on the surface of the protein, known as the...
Protein-ligand interactions are quite specific; even though numerous potential ligands surround a cellular protein at any given time, only a particular ligand can bind to that protein. Moreover, a ligand binds only to a dedicated area on the surface of the protein, known as the...
00:49Ligand Binding and Linkage
Allosteric proteins have more than one ligand binding site; the binding of a ligand to any of these sites influences the binding of ligands to the other sites. When a protein is allosteric, its binding sites are called coupled or linked. In the case of enzymes, the site that binds to the substrate is known as the active site and the other site is known as the regulatory site. When a ligand binds to the regulatory site, this leads to conformational changes in the protein that can influence the...
02:53Conservative Site-specific Recombination and Phase Variation
Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...
The recognition sites for Cre recombinase called LoxP...
01:19Allosteric Proteins-ATCase
Binding sites linkages can regulate a protein's function. For example, enzyme activity is often regulated through a feedback mechanism where the end product of the biochemical process serves as an inhibitor.
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis pathway,...
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis pathway,...