Crystal Structures of Arabidopsis thaliana Cell-Wall Invertase Mutants in Complex with Sucrose

Lammens, Willem; Roy, Katrien Le; Laere, André Van; Rabijns, A.; Ende, Wim Van den

doi:10.1016/j.jmb.2007.12.074

Cited by 70 publications

(86 citation statements)

References 41 publications

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“…The N/W couple is part of a flexible loop structure, comparable to the D/K couple, as observed in AtcwINV1 ( Fig. 7; Lammens et al, 2008). Intriguingly, all S-type enzymes (1-SSTs, 6-SFTs, and vacuolar invertases) contain a D and an R at these positions (Fig.…”

Section: Discussion Marked Differences Between S and F Enzyme Typesmentioning

confidence: 61%

“…Several three-dimensional structures have been unraveled within GH32 (for review, see Lammens et al, 2008), including a 1-FEH from chicory (Verhaest et al, 2005) and, most recently, a CWI from Arabidopsis (Verhaest et al, 2006).…”

Section: Parallels With Donor Substrate Selectivity In the Cwi/feh Groupmentioning

confidence: 99%

“…limited to the VI/FT subgroup, since no FT-type enzymes have been discovered in the CWI/FEH subgroup. According to Lammens et al (2008), so far all family GH32 enzymes bind the terminal Fru (from Suc or a fructan as the donor substrate) in exactly the same way at the 21 position, consisting of the active site formed by the catalytic triad (Fig. 2).…”

Section: Parallels With Donor Substrate Selectivity In the Cwi/feh Groupmentioning

confidence: 99%

“…By contrast, FEHs are believed to have evolved from cell wall invertases (CWIs) or from an ancestral b-fructosidase type of enzyme , 2008 capable of degrading both Suc and fructans. VIs, CWIs, FEHs, and FTs all belong to family 32 of glycoside hydrolases (GH32; Lammens et al, 2008).Among the FTs, two major groups can be further discerned: FTs that use Suc as a preferential donor substrate on the one hand (S-type enzymes: 1-SST and 6-SFT) and FTs that use fructans (and not Suc) as a preferential donor substrate on the other hand (F-type enzymes: 1-FFT and 6G-FFT). From a functional point of view, S-type enzymes are still more similar to the VIs from which they evolved, since both enzymes split Suc, a property that was lost or strongly reduced in F-type enzymes.…”

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confidence: 99%

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Transforming a Fructan:Fructan 6G-Fructosyltransferase from Perennial Ryegrass into a Sucrose:Sucrose 1-Fructosyltransferase

et al. 2008

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Fructosyltransferases (FTs) synthesize fructans, fructose polymers accumulating in economically important cool-season grasses and cereals. FTs might be crucial for plant survival under stress conditions in species in which fructans represent the major form of reserve carbohydrate, such as perennial ryegrass (Lolium perenne). Two FT types can be distinguished: those using sucrose (S-type enzymes: sucrose:sucrose 1-fructosyltransferase , sucrose:fructan 6-fructosyltransferase) and those using fructans (F-type enzymes: fructan:fructan 1-fructosyltransferase , fructan:fructan 6G-fructosyltransferase [6G-FFT]) as preferential donor substrate. Here, we report, to our knowledge for the first time, the transformation of an F-type enzyme (6G-FFT/1-FFT) into an S-type enzyme (1-SST) using perennial ryegrass 6G-FFT/1-FFT (Lp6G-FFT/1-FFT) and 1-SST (Lp1-SST) as model enzymes. This transformation was accomplished by mutating three amino acids (N340D, W343R, and S415N) in the vicinity of the active site of Lp6G-FFT/1-FFT. In addition, effects of each amino acid mutation alone or in combination have been studied. Our results strongly suggest that the amino acid at position 343 (tryptophan or arginine) can greatly determine the donor substrate characteristics by influencing the position of the amino acid at position 340. Moreover, the presence of arginine-343 negatively affects the formation of neofructan-type linkages. The results are compared with recent findings on donor substrate selectivity within the group of plant cell wall invertases and fructan exohydrolases. Taken together, these insights contribute to our knowledge of structure/function relationships within plant family 32 glycosyl hydrolases and open the way to the production of tailor-made fructans on a larger scale.Fructans are Fru polymers that can be considered as an extension of Suc, accumulating above a certain threshold Suc concentration (Cairns and Pollock, 1988;Guerrand et al., 1996;Maleux and Van den Ende, 2007). Fructans occur in many dicot and monocot plant species mainly belonging to Asteraceae, Liliaceae, and Poaceae (Hendry, 1993;Prud'homme et al., 2007;Shiomi et al., 2007;Van den Ende and Van Laere, 2007;Yoshida et al., 2007). Nowadays, research is mainly focused on economically important cereals (wheat [Triticum aestivum] and barley [Hordeum vulgare]) and forage grasses (mainly Lolium species). Depending on the linkage type between the fructosyl residues and the position of the Glc residue (Lewis, 1993), several fructan types can be distinguished. Fructans with a terminal Glc residue include the b(2-1)-type fructans (inulin, principally occurring in dicots) and the linear b(2-6) (levan)-or branched-type fructans (graminan) with both b(2-6) and b(2-1) linkages (as occurring in bacteria and cereals, respectively). Fructans with an internal Glc residue include the neoinulin and neolevan types (occurring in monocots such as Allium, Asparagus, and Lolium).Apart from their function as a vacuolar storage carbohydrate, fructans may help to protect plants ag...

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Section: Discussion Marked Differences Between S and F Enzyme Typesmentioning

confidence: 61%

Section: Parallels With Donor Substrate Selectivity In the Cwi/feh Groupmentioning

confidence: 99%

Section: Parallels With Donor Substrate Selectivity In the Cwi/feh Groupmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Transforming a Fructan:Fructan 6G-Fructosyltransferase from Perennial Ryegrass into a Sucrose:Sucrose 1-Fructosyltransferase

et al. 2008

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Structural and enzymatic analyses of Anabaena heterocyst‐specific alkaline invertase InvB

Xie

Cai

et al. 2018

FEBS Letters

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Anabaena sp. PCC 7120 encodes two alkaline/neutral invertases, namely InvA and InvB. Following our recently reported InvA structure, here we report the crystal structure of the heterocyst-specific InvB. Despite sharing an overall structure similar to InvA, InvB possesses a much higher catalytic activity. Structural comparisons of the catalytic pockets reveal that Arg430 of InvB adopts a different conformation, which may facilitate the deprotonation of the catalytic residue Glu415. We propose that the higher activity may be responsible for the vital role of InvB in heterocyst development and nitrogen fixation. Furthermore, phylogenetic analysis combined with activity assays also suggests the role of this highly conserved arginine in plants and cyanobacteria, as well as some proteobacteria living in highly extreme environments.

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Mutational analysis of conserved regions harboring catalytic triad residues of the levansucrase protein encoded by the lsc‐3 gene (lsc3) of Pseudomonas syringae pv. tomato DC3000

Mardo

Visnapuu

Vija

et al. 2013

Biotech and App Biochem

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Levansucrase encoded by the lsc-3 (lsc3) gene at genomic locus PSPTOA0032 of Pseudomonas syringae pv. tomato DC3000 was mutationally analyzed. Altogether, 18 single-amino-acid mutants of 13 positions of Lsc3 were studied for catalytic properties, including production of fructooligosaccharides (FOS). Asp62, Asp219, and Glu303 were proved as members of the catalytic triad. Respective alanine replacement mutants were practically inactive with their k cat values reduced up to ∼130,000 times. Additionally, the requirements of Trp61, Gln301, and Arg304, located in conserved sequence blocks around the catalytic triad positions for the catalysis were shown. The catalytic significance of the position equivalent to Arg304 was shown for levansucrases for the first time. Replacement of Gln301 specifically affected the polymerizing ability of Lsc3. The Gln301Ala mutant was largely hydrolytic and produced 31 times less FOS than the wild type. Despite high conservation grades, Leu66, Pro220, Asp225, and His306 tolerated replacement well. Quantification of produced FOS showed a high biotechnological potential of Lsc3. Using 1 mg of Lsc3 protein, 15.4 g of FOS with a degree of polymerization from 3 to 7 can be synthesized in a 20 H reaction with 1,200 mM sucrose. Our expression system allowed us to produce up to 30 mg of Lsc3 protein from 1 L of induced culture of recombinant Escherichia coli. C 2013

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Crystal Structures of Arabidopsis thaliana Cell-Wall Invertase Mutants in Complex with Sucrose

Cited by 70 publications

References 41 publications

Transforming a Fructan:Fructan 6G-Fructosyltransferase from Perennial Ryegrass into a Sucrose:Sucrose 1-Fructosyltransferase

Transforming a Fructan:Fructan 6G-Fructosyltransferase from Perennial Ryegrass into a Sucrose:Sucrose 1-Fructosyltransferase

Structural and enzymatic analyses of Anabaena heterocyst‐specific alkaline invertase InvB

Mutational analysis of conserved regions harboring catalytic triad residues of the levansucrase protein encoded by the lsc‐3 gene (lsc3) of Pseudomonas syringae pv. tomato DC3000

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