X-ray crystallographic study of xylopentaose binding toPseudomonas fluorescens xylanase A

Leggio, Leila Lo; Jenkins, John A.; Harris, Gillian W.; Pickersgill, Richard W.

doi:10.1002/1097-0134(20001115)41:3<362::aid-prot80>3.0.co;2-n

Cited by 31 publications

(33 citation statements)

References 45 publications

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“…The xylooligosacharide conformation obtained in this way keeps the reported typical xylan 3-fold helix pattern. Furthermore, the five central xylose units are in a conformation most similar to that experimentally observed in xylopentaose bound to Pseudomonas fluorescens xylanase A (PDB code 1EZN) (44).…”

Section: Methodssupporting

confidence: 79%

“…The recognition of the GlcA carboxylate must be critical, because soaking of crystals in nondecorated xylooligosaccharides failed in providing observ-able complexes. In addition, O1, O2, O3, O4, and O5 also make many polar interactions with the Xyn30D-CBM35 residues Glu 31 , Tyr 34 , Asn 44 , and Asn 132 , both directly and through several well ordered water molecules, which keeps the GlcA moiety in a very fix position common to the GlcA and aldouronic soaked crystals. On the contrary, the attached xylose unit does not make any direct interaction with residues from the Xyn30D-CBM35 domain, and, in agreement with this observation, the electron density map shows weaker density at this position precluding building additional xylose units.…”

Section: -Ser 537mentioning

confidence: 99%

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Structural Analysis of Glucuronoxylan-specific Xyn30D and Its Attached CBM35 Domain Gives Insights into the Role of Modularity in Specificity*

Sainz-Polo

Valenzuela

González

et al. 2014

Journal of Biological Chemistry

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Background: Xylanases are crucial in plant cell wall recycling. Results: A glucuronoxylan-specific xylanase is attached to its binding module with moderate flexibility. This CBM35 displays novel structural features regulating specificity. Conclusion: Depolymerization of highly substituted xylans and an oriented interaction with its target substrate are proposed. Significance: Unraveling the mechanisms ruling modularity is essential to understanding the biomass deconstruction and to producing efficient biocatalysts.

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Section: Methodssupporting

confidence: 79%

Section: -Ser 537mentioning

confidence: 99%

Structural Analysis of Glucuronoxylan-specific Xyn30D and Its Attached CBM35 Domain Gives Insights into the Role of Modularity in Specificity*

Sainz-Polo

Valenzuela

González

et al. 2014

Journal of Biological Chemistry

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“…9,10) It is known that several amino acid residues are widely conserved as well the catalytic amino acid residues, forming subsite À2 to þ1 in GH10 xylanases. [11][12][13] The roles of some of the amino acid residues around the active cleft have been determined by mutation analysis and structural analysis with substrate complex. [14][15][16][17][18] We noticed that an arginine residue is conserved near the proton donor glutamic acid residue in the middle of loop 4 following strand 4 in the three-dimensional structures of GH10 xylanases.…”

supporting

confidence: 55%

“…31) Roberge et al substituted the conserved tyrosine residue with phenylalanine in Streptomyces lividans xylanase A and concluded that this hydrogen bond did not appear to be important for the catalytic activity. 32) Furthermore, some GH10 xylanases naturally have phenylalanine residue at the position, 12) again indicating that the phenolic hydroxyl group is not essential for the activity. These results agree with our The enzyme reaction was performed at pH 6.6 (*) or pH 7.0 (#).…”

Section: Ph-activity Relationships Of the Mutant Xylanasesmentioning

confidence: 99%

The Role of Conserved Arginine Residue in Loop 4 of Glycoside Hydrolase Family 10 Xylanases

Nishimoto

Kitaoka

Fushinobu

et al. 2005

Bioscience, Biotechnology, and Biochemistry

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An arginine residue in loop 4 connecting strand 4 and -helix 4 is conserved in glycoside hydrolase family 10 (GH10) xylanases. The arginine residues, Arg 204 in xylanase A from Bacillus halodurans C-125 (XynA) and Arg 196 in xylanase B from Clostridium stercorarium F9 (XynB), were replaced by glutamic acid, lysine, or glutamine residues (XynA R204E, K and Q, and XynB R196E, K and Q). The pH-k cat =K m and the pH-k cat relationships of these mutant enzymes were measured. The pK e2 and pK es2 values calculated from these curves were 8.59 and 8.29 (R204E), 8.59 and 8.10 (R204K), 8.61 and 8.19 (R204Q), 7.42 and 7.19 (R196E), 7.49 and 7.18 (R196K), and 7.86 and 7.38 (R196Q) respectively. Only the pK es2 value of arginine derivatives was less than those of the wild types (8.49 and 9.39 [XynA] and 7.62 and 7.82 [XynB]). These results suggest that the conserved arginine residue in GH10 xylanases increases the pK a value of the proton donor Glu during substrate binding. The arginine residue is considered to clamp the proton donor and subsite þ1 to prevent structural change during substrate binding.

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“…At the Ϫ2 subsite the xylose interacts with residues that are also highly conserved in family 10 xylanases with the O-2 of the sugar hydrogen bonding to a glutamate and tryptophan, the O-3 to an asparagine, and the endocyclic oxygen to a lysine. There is a paucity of information, however, on the mechanism by which the aglycone region of the substrate binding cleft of these enzymes interacts with the xylan backbone, although the three-dimensional structure of Cellvibrio japonicus xylanase 10A (CjXyn10A) in complex with xylopentaose bound to subsites Ϫ1 to ϩ4 has been described (7,15). These studies showed that a highly conserved aromatic residue stacks against the xylose at the ϩ1 subsite, and although hydrophobic stacking interactions at the ϩ3 and ϩ4 subsites were the primary mechanism of protein-substrate recognition, these amino acids are not invariant in GH10 glycoside hydrolases, suggesting that xylan binding in the distal aglycone region of this enzyme family is variable.…”

mentioning

confidence: 99%

The Mechanisms by Which Family 10 Glycoside Hydrolases Bind Decorated Substrates

Pell

Taylor

Gloster

et al. 2004

Journal of Biological Chemistry

161

136

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Endo-␤-1,4-xylanases (xylanases), which cleave ␤-1,4 glycosidic bonds in the xylan backbone, are important components of the repertoire of enzymes that catalyze plant cell wall degradation. The mechanism by which these enzymes are able to hydrolyze a range of decorated xylans remains unclear. Here we reveal the threedimensional structure, determined by x-ray crystallography, and the catalytic properties of the Cellvibrio mixtus enzyme Xyn10B (CmXyn10B), the most active GH10 xylanase described to date. The crystal structure of the enzyme in complex with xylopentaose reveals that at the ؉1 subsite the xylose moiety is sandwiched between hydrophobic residues, which is likely to mediate tighter binding than in other GH10 xylanases. The crystal structure of the xylanase in complex with a range of decorated xylooligosaccharides reveals how this enzyme is able to hydrolyze substituted xylan. Solvent exposure of the O-2 groups of xylose at the ؉4, ؉3, ؉1, and ؊3 subsites may allow accommodation of the ␣-1,2-linked 4-O-methyl-D-glucuronic acid side chain in glucuronoxylan at these locations. Furthermore, the uronic acid makes hydrogen bonds and hydrophobic interactions with the enzyme at the ؉1 subsite, indicating that the sugar decorations in glucuronoxylan are targeted to this proximal aglycone binding site. Accommodation of 3-linked L-arabinofuranoside decorations is observed in the ؊2 subsite and could, most likely, be tolerated when bound to xylosides in ؊3 and ؉4. A notable feature of the binding mode of decorated substrates is the way in which the subsite specificities are tailored both to prevent the formation of "dead-end" reaction products and to facilitate synergy with the xylan degradation-accessory enzymes such as ␣-glucuronidase. The data described in this report and in the accompanying paper (Fujimoto, Z., Kaneko, S., Kuno, A., Kobayashi, H., Kusakabe, I., and Mizuno, H. (2004) J. Biol. Chem. 279, 9606 -9614) indicate that the complementarity in the binding of decorated substrates between the glycone and aglycone regions appears to be a conserved feature of GH10 xylanases.

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X-ray crystallographic study of xylopentaose binding toPseudomonas fluorescens xylanase A

Cited by 31 publications

References 45 publications

Structural Analysis of Glucuronoxylan-specific Xyn30D and Its Attached CBM35 Domain Gives Insights into the Role of Modularity in Specificity*

Structural Analysis of Glucuronoxylan-specific Xyn30D and Its Attached CBM35 Domain Gives Insights into the Role of Modularity in Specificity*

The Role of Conserved Arginine Residue in Loop 4 of Glycoside Hydrolase Family 10 Xylanases

The Mechanisms by Which Family 10 Glycoside Hydrolases Bind Decorated Substrates

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