X-ray diffraction structure of a cell-wall invertase from<i>Arabidopsis thaliana</i>

Verhaest, Maureen; Lammens, Willem; Roy, Katrien Le; Coninck, Barbara De; Ranter, C. J. De; Laere, André Van; Ende, Wim Van den; Rabijns, A.

doi:10.1107/s0907444906044489

Cited by 70 publications

(87 citation statements)

References 41 publications

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“…This result indicates that the purification procedure maintained the secondary native structure of the protein. In addition, the (Linde et al 2009) and from Arabidopsis thaliana (Verhaest et al 2006) have propeller and β-sheet motifs, as verified for the enzyme from A. phoenicis. These characteristics are found in many glycoside hydrolases from GH32 family, which includes β-D-fructofuranosidases.…”

Section: Circular Dichroismmentioning

confidence: 99%

Biochemical properties of an extracellular β-D-fructofuranosidase II produced by Aspergillus phoenicis under Solid-Sate Fermentation using soy bran as substrate

Rustiguel¹,

Oliveira²,

Terenzi³

et al. 2011

Electro Journal of Biotech

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The filamentous fungus A. phoenicis produced high levels of β-D-fructofuranosidase (FFase) when grown for 72 hrs under Solid-State Fermentation (SSF), using soy bran moistened with tap water (1:0.5 w/v) as substrate/carbon source. Two isoforms (I and II) were obtained, and FFase II was purified 18-fold to apparent homogeneity with 14% recovery. The native molecular mass of the glycoprotein (12% of carbohydrate content) was 158.5 kDa with two subunits of 85 kDa estimated by SDS-PAGE. Optima of temperature and pH were 55ºC and 4.5. The enzyme was stable for more than 1 hr at 50ºC and was also stable in a pH range from 7.0 to 8.0. FFase II retained 80% of activity after storage at 4ºC by 200 hrs. Dichroism analysis showed the presence of random and β-sheet structure. A. phoenicis

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Section: Circular Dichroismmentioning

confidence: 99%

Biochemical properties of an extracellular β-D-fructofuranosidase II produced by Aspergillus phoenicis under Solid-Sate Fermentation using soy bran as substrate

Rustiguel¹,

Oliveira²,

Terenzi³

et al. 2011

Electro Journal of Biotech

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“…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%

“…1). Moreover, a molecular modeling approach realized in silico (using Arabidopsis [Arabidopsis thaliana] cell wall invertase 1 [AtcwINV1] as a template; Verhaest et al, 2006; Protein Data Bank identifier 2AC1) allowed us to further reduce the number of promising candidates. The active site of all GH32 enzymes is characterized by three highly conserved amino acids in the WMNDPNG, RDP, and EC motifs (Reddy and Maley, 1996;Schroeven et al, 2008), and only the amino acids within a 15-Ǻ distance from the catalytic triad center were taken into account (Fig.…”

Section: Designing Mutants Based On Multiple Sequence Alignments and mentioning

confidence: 99%

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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“…Homology models of the native Nin88 were built in modeller9v7 (Eswar et al, 2006) with multiple template methods using the known protein crystal structures of AtcwINV1 (Protein Data Bank [PDB] code 2AC1; Verhaest et al, 2006) and 1-FEH IIa of chicory (Cichorium intybus; PDB code 1ST8; Verhaest et al, 2005) as model-building templates. The models were further optimized by subjecting them to the computer program Brugel (Delhaise et al, 1984).…”

Section: Homology Modeling Of Nin88mentioning

confidence: 99%

Understanding the Role of Defective Invertases in Plants: Tobacco Nin88 Fails to Degrade Sucrose

et al. 2013

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Cell wall invertases (cwINVs), with a high affinity for the cell wall, are fundamental enzymes in the control of plant growth, development, and carbon partitioning. Most interestingly, defective cwINVs have been described in several plant species. Their highly attenuated sucrose (Suc)-hydrolyzing capacity is due to the absence of aspartate-239 (Asp-239) and tryptophan-47 (Trp-47) homologs, crucial players for stable binding in the active site and subsequent hydrolysis. However, so far, the precise roles of such defective cwINVs remain unclear. In this paper, we report on the functional characterization of tobacco (Nicotiana tabacum) Nin88, a presumed fully active cwINV playing a crucial role during pollen development. It is demonstrated here that Nin88, lacking both Asp-239 and Trp-47 homologs, has no invertase activity. This was further supported by modeling studies and site-directed mutagenesis experiments, introducing both Asp-239 and Trp-47 homologs, leading to an enzyme with a distinct Suc-hydrolyzing capacity. In vitro experiments suggest that the addition of Nin88 counteracts the unproductive and rather aspecific binding of tobacco cwINV1 to the wall, leading to higher activities in the presence of Suc and a more efficient interaction with its cell wall inhibitor. A working model is presented based on these findings, allowing speculation on the putative role of Nin88 in muro. The results presented in this work are an important first step toward unraveling the specific roles of plant defective cwINVs.

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X-ray diffraction structure of a cell-wall invertase fromArabidopsis thaliana

Cited by 70 publications

References 41 publications

Biochemical properties of an extracellular β-D-fructofuranosidase II produced by Aspergillus phoenicis under Solid-Sate Fermentation using soy bran as substrate

Biochemical properties of an extracellular β-D-fructofuranosidase II produced by Aspergillus phoenicis under Solid-Sate Fermentation using soy bran as substrate

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

Understanding the Role of Defective Invertases in Plants: Tobacco Nin88 Fails to Degrade Sucrose

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