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Exploring natural genetic variation in tomato sucrose synthases on the basis of increased kinetic properties.

ABSTRACT: Sucrose synthase (SuSy) is one key enzyme directly hydrolyzing sucrose to supply substrates for plant metabolism, and is considered to be a biomarker for plant sink strength. Improvement in plant sink strength could lead to enhanced plant growth and yield. Cultivated tomatoes are known to have a narrow genetic diversity, which hampers further breeding for novel and improved traits in new cultivars. In this study, we observed limited genetic variation in SuSy1, SuSy3 and SuSy4 in 53 accessions of cultivated tomato and landraces, but identified a wealth of genetic diversity in 32 accessions of related wild species. The variation in the deduced amino acid sequences was grouped into 23, 22, and 17 distinct haplotypes for SuSy1/3/4, respectively. Strikingly, all known substrate binding sites were highly conserved, as well as most of the phosphorylation sites except in SuSy1. Two SuSy1 and three SuSy3 protein variants were heterologously expressed to study the effect of the amino acid changes on enzyme kinetic properties, i.e. maximal sucrose hydrolyzing capacity (Vmax), affinity for sucrose (Km), and catalytic efficiency (Vmax/Km) at 25°C and 16°C. SuSy1-haplotype#3 containing phosphorylation site Ser-16 did not have an improvement in the kinetic properties compared to the reference SuSy1-haplotype#1 containing Arg-16. Meanwhile SuSy3-haplotype#9 from a wild accession, containing four amino acid changes S53A, S106I, E727D and K741E, showed an increase in Vmax/Km at 16°C compared to the reference SuSy3-haplotype#1. This study demonstrates that SuSy kinetic properties can be enhanced by exploiting natural variation, and the potential of this enzyme to improve sucrose metabolism and eventually sink strength in planta.

SUBMITTER: Dinh QD

PROVIDER: S-EPMC6205638 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Exploring natural genetic variation in tomato sucrose synthases on the basis of increased kinetic properties.

Dinh Quy-Dung QD Finkers Richard R Westphal Adrie H AH van Dongen Walter M A M WMAM Visser Richard G F RGF Trindade Luisa M LM

PloS one 20181029 10

Sucrose synthase (SuSy) is one key enzyme directly hydrolyzing sucrose to supply substrates for plant metabolism, and is considered to be a biomarker for plant sink strength. Improvement in plant sink strength could lead to enhanced plant growth and yield. Cultivated tomatoes are known to have a narrow genetic diversity, which hampers further breeding for novel and improved traits in new cultivars. In this study, we observed limited genetic variation in SuSy1, SuSy3 and SuSy4 in 53 accessions of ...[more]

PMID: 30372500

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Project description:Sucrose is the end product of photosynthesis and the primary sugar transported in the phloem of most plants. Sucrose synthase (SuSy) is a glycosyl transferase enzyme that plays a key role in sugar metabolism, primarily in sink tissues. SuSy catalyzes the reversible cleavage of sucrose into fructose and either uridine diphosphate glucose (UDP-G) or adenosine diphosphate glucose (ADP-G). The products of sucrose cleavage by SuSy are available for many metabolic pathways, such as energy production, primary-metabolite production, and the synthesis of complex carbohydrates. SuSy proteins are usually homotetramers with an average monomeric molecular weight of about 90 kD (about 800 amino acids long). Plant SuSy isozymes are mainly located in the cytosol or adjacent to plasma membrane, but some SuSy proteins are found in the cell wall, vacuoles, and mitochondria. Plant SUS gene families are usually small, containing between four to seven genes, with distinct exon-intron structures. Plant SUS genes are divided into three separate clades, which are present in both monocots and dicots. A comprehensive phylogenetic analysis indicates that a first SUS duplication event may have occurred before the divergence of the gymnosperms and angiosperms and a second duplication event probably occurred in a common angiosperm ancestor, leading to the existence of all three clades in both monocots and dicots. Plants with reduced SuSy activity have been shown to have reduced growth, reduced starch, cellulose or callose synthesis, reduced tolerance to anaerobic-stress conditions and altered shoot apical meristem function and leaf morphology. Plants overexpressing SUS have shown increased growth, increased xylem area and xylem cell-wall width, and increased cellulose and starch contents, making SUS high-potential candidate genes for the improvement of agricultural traits in crop plants. This review summarizes the current knowledge regarding plant SuSy, including newly discovered possible developmental roles for SuSy in meristem functioning that involve sugar and hormonal signaling.

Project description:Polyhydroxyalkanoates (PHAs) are biopolyesters composed of hydroxy fatty acids, which represent a complex class of storage polyesters. They are synthesized by a wide range of different Gram-positive and Gram-negative bacteria, as well as by some Archaea, and are deposited as insoluble cytoplasmic inclusions. Polyester synthases are the key enzymes of polyester biosynthesis and catalyse the conversion of (R)-hydroxyacyl-CoA thioesters to polyesters with the concomitant release of CoA. These soluble enzymes turn into amphipathic enzymes upon covalent catalysis of polyester-chain formation. A self-assembly process is initiated resulting in the formation of insoluble cytoplasmic inclusions with a phospholipid monolayer and covalently attached polyester synthases at the surface. Surface-attached polyester synthases show a marked increase in enzyme activity. These polyester synthases have only recently been biochemically characterized. An overview of these recent findings is provided. At present, 59 polyester synthase structural genes from 45 different bacteria have been cloned and the nucleotide sequences have been obtained. The multiple alignment of the primary structures of these polyester synthases show an overall identity of 8-96% with only eight strictly conserved amino acid residues. Polyester synthases can been assigned to four classes based on their substrate specificity and subunit composition. The current knowledge on the organization of the polyester synthase genes, and other genes encoding proteins related to PHA metabolism, is compiled. In addition, the primary structures of the 59 PHA synthases are aligned and analysed with respect to highly conserved amino acids, and biochemical features of polyester synthases are described. The proposed catalytic mechanism based on similarities to alpha/beta-hydrolases and mutational analysis is discussed. Different threading algorithms suggest that polyester synthases belong to the alpha/beta-hydrolase superfamily, with a conserved cysteine residue as catalytic nucleophile. This review provides a survey of the known biochemical features of these unique enzymes and their proposed catalytic mechanism.

Dataset Information

Exploring natural genetic variation in tomato sucrose synthases on the basis of increased kinetic properties.

Publications

Exploring natural genetic variation in tomato sucrose synthases on the basis of increased kinetic properties.

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