Three-dimensional structure of human lysosomal aspartylglucosaminidase

Oinonen, Carita; Tikkanen, Ritva; Rouvinen, Juha; Peltonen, Leena

doi:10.1038/nsb1295-1102

Cited by 172 publications

(199 citation statements)

References 32 publications

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“…The GGT heterodimer has a stacked ␣␤␤␣-core; the ␤-strands of the L and S subunits form the two central ␤-sheets, which are sandwiched by ␣-helices. The topology of GGT is similar to those observed for Ntn-hydrolase superfamily members such as aspartylglucosaminidase (22) and penicillin acylase (23). The ␤-strands comprising the two central ␤-sheets and nearby ␣-helices are topologically conserved among the members of this superfamily; the main-chain atoms of the 66 residues comprising the core of E. coli GGT are superimposable on those of human aspartylglucosaminidase with a rms deviation of 1.3 Å.…”

Section: Resultsmentioning

confidence: 59%

Crystal structures of γ-glutamyltranspeptidase from Escherichia coli , a key enzyme in glutathione metabolism, and its reaction intermediate

Okada

Suzuki

Wada

et al. 2006

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

159

211

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␥-Glutamyltranspeptidase (GGT) is a heterodimic enzyme that is generated from the precursor protein through posttranslational processing and catalyzes the hydrolysis of ␥-glutamyl bonds in ␥-glutamyl compounds such as glutathione and͞or the transfer of the ␥-glutamyl group to other amino acids and peptides. We have determined the crystal structure of GGT from Escherichia coli K-12 at 1.95 Å resolution. GGT has a stacked ␣␤␤␣ fold comprising the large and small subunits, similar to the folds seen in members of the N-terminal nucleophile hydrolase superfamily. The active site Thr-391, the N-terminal residue of the small subunit, is located in the groove, from which the pocket for ␥-glutamyl moiety binding follows. We have further determined the structure of the ␥-glutamyl

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Section: Resultsmentioning

confidence: 59%

Crystal structures of γ-glutamyltranspeptidase from Escherichia coli , a key enzyme in glutathione metabolism, and its reaction intermediate

Okada

Suzuki

Wada

et al. 2006

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

159

211

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“…The funnel is formed by loops that connect the two P-sheets and by loops connecting the smaller P-sheet and a-helices associated with it. Oinonen et al (1995) published the structure of human glycosylasparaginase in free form and as a complex with aspartic acid. All residues that bind aspartic acid in the human enzyme are contributed by two loops, P-strand B2 to p-strand B3, and P-strand B4 to a-helix B 1.…”

Section: Resultsmentioning

confidence: 99%

“…This is primarily caused by the changes in the conformations of the loops between a-helix A2 and p-strand A2 and between P-strand B6 and p-strand B7, which are among the loops that differ most between the two enzymes, and by differences in some residues at the edge of the cleft, most notably the substitution of Gln 254 in the bacterial enzyme for Phe 278 in the human enzyme. The latter substitution is significant because Oinonen et al (1995) suggested that Trp 11 and Phe 278 may be important for oligosaccharide binding. Although Trp 11 is conserved, the substitution of Gln for Phe 278 does not preclude this observation because both aromatic and polar-neutral residues are equally suitable for oligosaccharide binding, although with possible differences in specificities.…”

Section: Resultsmentioning

confidence: 99%

Crystal structure of glycosylasparaginase from Flavobacterium meningosepticum

et al. 1998

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The crystal structure of recombinant glycosylasparaginase from Flavobacterium meningosepticum has been determined at 2.32 a resolution. This enzyme is aglycoamidase that cleaves the link between the asparagine and the N-acetylglucosamine of N-linked oligosaccharides and plays a major role in the degradation of glycoproteins. The three-dimensional structure of the bacterial enzyme is very similar to that of the human enzyme, although it lacks the four disulfide bridges found in the human enzyme. The main difference is the absence of a small random coil domain at the end of the a-chain that forms part of the substrate binding cleft and that has a role in the stabilization of the tetramer of the human enzyme. The bacterial glycosylasparaginase is observed as an (a/3)2-tetramer in the crystal, despite being a dimer in solution. The study of the structure of the bacterial enzyme allows further evaluation of the effects of disease-causing mutations in the human enzyme and confirms the suitability of the bacterial enzyme as a model for functional analysis.

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“…Bacterial L-asparaginases were the first threonine amidohydrolases described in the literature. Only recently, two other threonine amidohydrolases, 20S proteasome [14] and aspartylglucosaminidase [15], have been reported. Both enzymes belong to the nonrelated superfamily of N-terminal nucleophile (Ntn) hydrolases [16].…”

mentioning

confidence: 99%

A covalently bound catalytic intermediate in Escherichia coli asparaginase : Crystal structure of a Thr‐89‐Val mutant

Palm¹,

Łubkowski²,

Derst³

et al. 1996

FEBS Letters

112

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Escherichia coli asparaginase |I catalyzes the hydrol-) sis of L-asparagine to L-aspartate via a threonine-bound acylenzyme intermediate. A nearly inactive mutant in which one of the active site threonines, Thr-89, was replaced by valine was constructed, expressed, and crystallized. Its structure, solved at 2.2 A resolution, shows high overall similarity to the wild-type enzyme, but an aspartyl moiety is covalently bound to Thr-12, resembling a reaction intermediate. Kinetic analysis confirms the deacylation deficiency, which is also explained on a structural basis. The previously identified oxyanion hole is described in more detail.:£ey words'." Asparaginase II; Acyl-enzyme intermediate; !'hreonine amidohydrolase; Enzymatic mechanism !. IntroductionAsparaginases catalyze the hydrolysis of L-asparagine to L~spartate and ammonia. The enzymes isolated from Escheri-'hia coli (EcA) and Erwinia chrysanthemi (ERA) have been ltilized as anti-leukemia drugs for many years [1]. Treatment vith asparaginases is often accompanied by severe side effects, vhich are partially attributed to the glutaminase activity of hese enzymes [2]. In order to understand the enzyme specifi-:ity and to ultimately eliminate the glutaminase activity, the nechanism of action must be elucidated. Several members of t larger family of homologous L-asparaginases have thus been horoughly investigated over many years.Results of kinetic measurements [3,4] indicated that the en-,ymatic reaction proceeds via a covalent intermediate, probtbly a ~-aspartyl enzyme (Fig. 1). This mechanism was con-,irmed by NMR studies with aspartate through the oxygen ~xchange reaction with bulk solvent at the side chain carboxdate [5]. These experiments showed that at pH < 5 L-aspartate ~ith a protonated side chain binds to the enzyme as tightly as ~-asparagine and it can also form an acyl-enzyme intermediate, subsequently hydrolyzed to L-aspartate. Thus, at low pH 3oth L-aspartate and L-asparagine can function as substrates. l'he nature and identity of the primary nucleophile of the mzyme participating in the formation of the acyl-enzyme in-*Corresponding author. Fax: (1) . Each has the same tetrameric quaternary structure as it has in solution, a homotetramer of approx. 4x330 residues. The tetramer is more accurately described as a dimer of dimers. Each of two identical active sites in each dimer is formed by both monomers. In the structure the active sites have been observed with and without aspartate as ligand. Part of the active site is covered by a flexible loop that contains the two important residues Thr-12 and Tyr-25. The hydroxyl groups of Thr-12 and Thr-89 are closest to the side chain carboxylate of an aspartate bound in the active site. These residues are the most likely candidates for the primary nucleophile. Bacterial L-asparaginases were the first threonine amidohydrolases described in the literature. Only recently, two other threonine amidohydrolases, 20S proteasome [14] and aspartylglucosaminidase [15], have been reported. Both enzymes belong t...

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Three-dimensional structure of human lysosomal aspartylglucosaminidase

Cited by 172 publications

References 32 publications

Crystal structures of γ-glutamyltranspeptidase from Escherichia coli , a key enzyme in glutathione metabolism, and its reaction intermediate

Crystal structures of γ-glutamyltranspeptidase from Escherichia coli , a key enzyme in glutathione metabolism, and its reaction intermediate

Crystal structure of glycosylasparaginase from Flavobacterium meningosepticum

A covalently bound catalytic intermediate in Escherichia coli asparaginase : Crystal structure of a Thr‐89‐Val mutant

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