Catalytic Versatility, Stability, and Evolution of the (βα)<sub>8</sub>-Barrel Enzyme Fold

Sterner, Reinhard; Höcker, Birte

doi:10.1021/cr030191z

Cited by 187 publications

(146 citation statements)

References 158 publications

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“…Moreover, we have confirmed that residues Lys-81, Asp-24, and Asp-102, in the vicinity of P1, are necessary for catalysis, because mutation to Ala resulted in an enzyme without detectable PLP synthase activity (data not shown). These data together establish the Pdx1 active site around P1 on the catalytic face of the (␤␣) 8 -barrel, as is consistent with placement of active sites for all structurally known enzymes of this fold (18). Furthermore, it is interesting that residues 47-56, which form the small helical segment ␣2Ј, are observed only in the ternary complex (Figs.…”

Section: Discussionsupporting

confidence: 80%

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Structure of a bacterial pyridoxal 5′-phosphate synthase complex

Strohmeier

Raschle

Mazurkiewicz

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

113

272

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Vitamin B6 is an essential metabolic cofactor that has more functions in humans than any other single nutrient. Its de novo biosynthesis occurs through two mutually exclusive pathways that are absent in animals. The predominant pathway found in most prokaryotes, fungi, and plants has only recently been discovered. It is distinguished by a glutamine amidotransferase, which is remarkable in that it alone can synthesize the cofactor form, pyridoxal 5-phosphate (PLP), directly from a triose and a pentose saccharide and glutamine. Here we report the 3D structure of the PLP synthase complex with substrate glutamine bound as well as those of the individual synthase and glutaminase subunits Pdx1 and Pdx2, respectively. The complex is made up of 24 protein units assembled like a cogwheel, a dodecameric Pdx1 to which 12 Pdx2 subunits attach. In contrast to the architecture of previously determined glutamine amidotransferases, macromolecular assembly is directed by an N-terminal ␣-helix on the synthase. Interaction with the synthase subunit leads to glutaminase activation, resulting in formation of an oxyanion hole, a prerequisite for catalysis. Mutagenesis permitted identification of the remote glutaminase and synthase catalytic centers and led us to propose a mechanism whereby ammonia shuttles between these active sites through a methionine-rich hydrophobic tunnel.3D structure ͉ ammonia tunnel ͉ glutamine amidotransferase ͉ oxyanion ͉ vitamin B6

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Section: Discussionsupporting

confidence: 80%

“…1e), also determined at 2.1-Å resolution (R work 14.4%; R free 19.4%; see SI Table 2). Pdx1 folds as a (␤␣) 8 -barrel (18), whose axis is oriented nearly perpendicular to the ring perimeter. A bulge created by the helix ␣8ЈЈ at the C terminus running parallel to helix ␣8 establishes the contact between adjacent (␤␣) 8 -barrels.…”

mentioning

confidence: 99%

Structure of a bacterial pyridoxal 5′-phosphate synthase complex

Strohmeier

Raschle

Mazurkiewicz

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

113

272

View full text Add to dashboard Cite

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“…Furthermore, as is seen in the figure, the segment 422-429 is located between the end of helices ␣1 and ␣8 in a kind of barrel "closure." This is a remarkable trait taking into account that the short loops located at the opposite site of the active site cavity are generally considered to be essential in keeping the barrel stability; in fact, this side of the barrel has been termed the "stability face" (27). This insertion is probably increasing the overall stability of ScAGal, and it might be a reason that would explain the heat stability observed for this enzyme (data not shown) and why it remains folded when performing SDS-PAGE in mild conditions (Fig.…”

Section: Resultsmentioning

confidence: 99%

Structural Analysis of Saccharomyces cerevisiae α-Galactosidase and Its Complexes with Natural Substrates Reveals New Insights into Substrate Specificity of GH27 Glycosidases

Fernández-Leiro

Pereira-Rodríguez

Cerdán

et al. 2010

Journal of Biological Chemistry

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␣-Galactosidases catalyze the hydrolysis of terminal ␣-1,6-galactosyl units from galacto-oligosaccharides and polymeric galactomannans. The crystal structures of tetrameric Saccharomyces cerevisiae ␣-galactosidase and its complexes with the substrates melibiose and raffinose have been determined to 1.95, 2.40, and 2.70 Å resolution. The monomer folds into a catalytic (␣/␤) 8 barrel and a C-terminal ␤-sandwich domain with unassigned function. This pattern is conserved with other family 27 glycosidases, but this enzyme presents a unique 45-residue insertion in the ␤-sandwich domain that folds over the barrel protecting it from the solvent and likely explaining its high stability. The structure of the complexes and the mutational analysis show that oligomerization is a key factor in substrate binding, as the substrates are located in a deep cavity making direct interactions with the adjacent subunit. Furthermore, docking analysis suggests that the supplementary domain could be involved in binding sugar units distal from the scissile bond, therefore ascribing a role in fine-tuning substrate specificity to this domain. It may also have a role in promoting association with the polymeric substrate because of the ordered arrangement that the four domains present in one face of the tetramer. Our analysis extends to other family 27 glycosidases, where some traits regarding specificity and oligomerization can be formulated on the basis of their sequence and the structures available. These results improve our knowledge on the activity of this important family of enzymes and give a deeper insight into the structural features that rule modularity and protein-carbohydrate interactions.Galactose is present in the oligosaccharides of many plant seeds and is also essential in structures as the hemicelluloses. These polymers build up the plant cell wall and represent a huge storage of carbon within the biosphere and might be an important source of renewable energy (1). In humans, mutations of the ␣-galactosidase gene cause incomplete degradation of glycolipids and glycoproteins, resulting in Fabry disease (2). Different strategies involving recombinant ␣-galactosidases are being developed for the treatment of this disease. Another interesting application is the conversion between the ABO blood groups, determined by differences in polysaccharide structures present in the surface of red blood cells. Some ␣-galactosidases are able to remove the ␣-linked terminal galactose that differs between O antigen (universal blood) and B antigen, and processes involving plant ␣-galactosidases are being developed to obtain O-type blood from B-type donors (3). Furthermore, the activity of ␣-galactosidase is of great interest in many biotechnological applications. It is used to improve the quality and yield of sucrose in the sugar beet industry by achieving an efficient raffinose and other galacto-oligosaccharide hydrolyses. In addition, the processing of soybean-related products and other legume-derived food with this enzyme reduces the content of...

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“…Unusual Deformations Found in TIM Barrel Scaffold-The TIM barrel is the most common protein fold and is adopted by about 10% of proteins with known three-dimensional structures (64). The functional versatility of TIM barrel proteins is generally based on the stability of the barrel architecture as well as the variability of the ␤␣-loops at the C-terminal ends of the ␤-strands ("catalytic face").…”

Section: Discussionmentioning

confidence: 99%

The Crystal Structure of Galacto-N-biose/Lacto-N-biose I Phosphorylase

Hidaka

Nishimoto

Kitaoka

et al. 2009

Journal of Biological Chemistry

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. GLNBP homologues are often found in human pathogenic and commensal bacteria, and their substrate specificities potentially define the nutritional acquisition ability of these microbes in their habitat. We report the crystal structures of GLNBP in five different ligand-binding forms. This is the first three-dimensional structure of glycoside hydrolase (GH) family 112. The GlcNAc-and GalNAc-bound forms provide structural insights into distinct substrate preferences of GLNBP and its homologues from pathogens. The catalytic domain consists of a partially broken TIM barrel fold that is structurally similar to a thermophilic ␤-galactosidase, strongly supporting the current classification of GLNBP homologues as one of the GH families. Anion binding induces a large conformational change by rotating a half-unit of the barrel. This is an unusual example of molecular adaptation of a TIM barrel scaffold to substrates.

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Catalytic Versatility, Stability, and Evolution of the (βα)₈-Barrel Enzyme Fold

Cited by 187 publications

References 158 publications

Structure of a bacterial pyridoxal 5′-phosphate synthase complex

Structure of a bacterial pyridoxal 5′-phosphate synthase complex

Structural Analysis of Saccharomyces cerevisiae α-Galactosidase and Its Complexes with Natural Substrates Reveals New Insights into Substrate Specificity of GH27 Glycosidases

The Crystal Structure of Galacto-N-biose/Lacto-N-biose I Phosphorylase

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Catalytic Versatility, Stability, and Evolution of the (βα)8-Barrel Enzyme Fold

Cited by 187 publications

References 158 publications

Structure of a bacterial pyridoxal 5′-phosphate synthase complex

Structure of a bacterial pyridoxal 5′-phosphate synthase complex

Structural Analysis of Saccharomyces cerevisiae α-Galactosidase and Its Complexes with Natural Substrates Reveals New Insights into Substrate Specificity of GH27 Glycosidases

The Crystal Structure of Galacto-N-biose/Lacto-N-biose I Phosphorylase

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Catalytic Versatility, Stability, and Evolution of the (βα)₈-Barrel Enzyme Fold