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ABSTRACT: Fructans are the main storage carbohydrates of temperate grasses, sustaining regrowth immediately after defoliation, as well as contributing to the nutritive value of feed. Fructan metabolism is based on the substrate sucrose and involves fructosyltransferases (FTs) for biosynthesis and fructan exohydrolases (FEHs) for degradation. Sucrose is also utilized by invertases (INVs), which hydrolyse it into its constituent monosaccharides for use in metabolism. The isolation, molecular characterization, functional analysis, and phylogenetic relationships of genes encoding FTs, FEHs, and INVs from temperate grasses are reviewed, with an emphasis on perennial ryegrass (Lolium perenne L.). The roles these enzymes play in fructan accumulation and remobilization, and future biotechnological applications in molecular plant breeding are discussed.
Plant Biotechnology Journal 10/2005; 3(5):459-74. · 5.44 Impact Factor
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ABSTRACT: Invasive forms of apicomplexan parasites contain secretory organelles called rhoptries that are essential for entry into host cells. We present a detailed characterization of an unusual rhoptry protein of the human malaria parasite Plasmodium falciparum, the rhoptry-associated membrane antigen (RAMA) that appears to have roles in both rhoptry biogenesis and host cell invasion. RAMA is synthesized as a 170-kDa protein in early trophozoites, several hours before rhoptry formation and is transiently localized within the endoplasmic reticulum and Golgi within lipid-rich microdomains. Regions of the Golgi membrane containing RAMA bud to form vesicles that later mature into rhoptries in a process that is inhibitable by brefeldin A. Other rhoptry proteins such as RhopH3 and RAP1 are found in close apposition with RAMA suggesting direct protein-protein interactions. We suggest that RAMA is involved in trafficking of these proteins into rhoptries. In rhoptries, RAMA is proteolytically processed to give a 60-kDa form that is anchored in the inner face of the rhoptry membrane by means of the glycosylphosphatidylinositol anchor. The p60 RAMA form is discharged from the rhoptries of free merozoites and binds to the red blood cell membrane by its most C-terminal region. In early ring stages RAMA is found in association with the parasitophorous vacuole.
Journal of Biological Chemistry 03/2004; 279(6):4648-56. · 4.77 Impact Factor
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ABSTRACT: A sucrose: sucrose 1-fructosyltransferase (1-SST) gene and cDNA (Lp 1-SST) from perennial ryegrass (Lolium perenne) were isolated. The Lp 1-SST gene was fully sequenced and shown to contain three exons and two introns. Nucleotide sequence analysis of the 4824 bp Lp 1-SST genomic sequence revealed 1618 bp of 5' UTR and an open reading frame of 1962 bp encoding a protein of 653 amino acids. Lp 1-SST is 95% identical to the tall fescue 1-SST and contains plant fructosyltransferase functional domains. Lp 1-SST corresponds to a single copy gene in perennial ryegrass, and is expressed in young leaf bases and mature leaf sheaths. The recombinant Lp 1-SST protein from corresponding cDNA expression in Pichia pastoris showed 1-SST activity.
Journal of Plant Physiology 12/2003; 160(11):1385-91. · 2.79 Impact Factor
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ABSTRACT: An invertase (LpFT2) cDNA from perennial ryegrass was isolated and sequenced. Nucleotide sequence analysis revealed an ORF of 2016 bp encoding a protein of 671 amino acids. LpFT2 is 76% identical to sugarcane soluble acid invertase, and contains invertase and fructosyltransferase functional domains. LpFT2 is present as a single copy gene and maps to the distal region of LG6 in perennial ryegrass. The expression pattern analysis of LpFT2 revealed transcript accumulation in seedlings and in mature leaf sheaths. The LpFT2 recombinant protein expressed in yeast showed invertase and fructan exohydrolase-like activities with complete breakdown of sucrose, 1-kestose (DP3), 1,1-kestotetraose (DP4) and 1,1,1-kestopentaose (DP5) into glucose and fructose.
Journal of Plant Physiology 09/2003; 160(8):903-11. · 2.79 Impact Factor
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ABSTRACT: Summary4-Coumarate : CoA-ligase (4CL, EC 6.2.1.12) catalyses the conversion of hydroxycinnamic acids to hydroxycinnamoyl-CoA thioesters, precursors for a variety of phenylpropanoid biosynthetic derivatives including lignins, flavonoids and phytoalexins. Three 4CL homologue cDNAs were isolated from a perennial ryegrass cDNA library. Sequence analysis indicates that Lp4CL2 and Lp4CL3 correspond to class I type 4CLs, while Lp4CL1 is most related to class II type 4CLs. Southern hybridisation analysis suggests that Lp4CL1, Lp4CL2 and Lp4CL3 each belong to a small multigene family. Northern hybridisation analysis revealed constitutive expression of the Lp4CL genes in perennial ryegrass. Lp4CL1 and Lp4CL3 were mapped to perennial ryegrass linkage groups 2 and 6 respectively.
Journal of Plant Physiology. 159(7):773-779.
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ABSTRACT: SummaryCinnamoyl-CoA reductase (CCR, EC 1.2.1.44) catalyses the conversion of hydroxycinnamoyl-CoA thioesters to cinnamaldehydes at the entry point to the monolignol-specific branch of the lignin biosynthetic pathway. A gene encoding CCR in perennial ryegrass (LpCCR1) was isolated and fully sequenced. LpCCR1 contains five exons of similar sizes to other CCRs and four introns in conserved positions. The 5′ untranslated region contains motifs common to lignin biosynthetic genes. LpCCR1 is present as a single copy gene and maps to LG7 in perennial ryegrass. LpCCR1 is constitutively expressed, is stimulated by mechanical wounding, and transcript abundance in stems increases with maturity.
Journal of Plant Physiology.
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ABSTRACT: SummaryThree cinnamyl alcohol dehydrogenase (CAD, EC 1.1.1.195) homologue cDNAs (LpCAD1, LpCAD2 and LpCAD3) were isolated and characterised in perennial ryegrass (Lolium perenne). Sequence analysis revealed a partial LpCAD3 clone is highly homologous to typical monocot CAD cDNAs while full-length LpCAD1 and LpCAD2 cDNAs show lower homology to known CAD cDNAs. Northern hybridisation analysis revealed LpCAD1 and LpCAD3 transcripts in lignifying tissues while LpCAD2 was uniquely abundant in stem tissue. All three CAD cDNAs are upregulated in response to mechanical wounding stimulus. Southern hybridisation analysis suggested that LpCAD3 is a single copy gene within the Lolium genome while LpCAD1 and LpCAD2 each belong to a small gene family.
Journal of Plant Physiology.
