Glycosylasparaginase-catalyzed Synthesis and Hydrolysis of β-Aspartyl Peptides

Tiina Noronkoski,Ivanka B Stoineva,Ivailo P Ivanov,Dimiter D Petkov,Ilkka Mononen

doi:10.1074/jbc.273.41.26295

Tiina Noronkoski, Ivanka B Stoineva + Show 3 more

Open Access

https://doi.org/10.1074/jbc.273.41.26295

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Abstract

beta-Aspartyl di- and tripeptides are common constituents of mammalian metabolism, but their formation and catabolism are not fully understood. In this study we provide evidence that glycosylasparaginase (aspartylglucosaminidase), an N-terminal nucleophile hydrolase involved in the hydrolysis of the N-glycosidic bond in glycoproteins, catalyzes the hydrolysis of beta-aspartyl peptides to form L-aspartic acid and amino acids or peptides. The enzyme also effectively catalyzes the synthesis of beta-aspartyl peptides by transferring the beta-aspartyl moiety from other beta-aspartyl peptides or beta-aspartylglycosylamine to a variety of amino acids and peptides. Furthermore, the enzyme can use L-asparagine as the beta-aspartyl donor in the formation of beta-aspartyl peptides. The data show that synthesis and degradation of beta-aspartyl peptides are new, significant functions of glycosylasparaginase and suggest that the enzyme could have an important role in the metabolism of beta-aspartyl peptides.

Highlights

␤-Aspartyl and ␥-glutamyl peptides are normal constituents of mammalian urine [1, 2] and tissues [3]
In the reaction Asn acted as ␤-aspartyl donor and serineamide as ␤-aspartyl acceptor in a kinetically controlled synthesis that closely resembles the glycosylasparaginase-catalyzed synthesis of the N-glycosidic bond by ␤-aspartylation of glycosylamines [18]. ␤-Aspartylglucosamine and ␤-aspartame can function as the ␤-aspartyl donor as demonstrated by the formation of ␤-aspartylserineamide in the presence of serineamide as an added nucleophile (Fig. 1)
Because the reaction components by themselves are virtually unreactive and no formation of ␤-aspartyl peptides was detected in the absence of glycosylasparaginase, the oligosaccharidyl-binding site in the enzyme must interact with the acceptor nucleophiles in the transamidase and transpeptidase reactions

Summary

Introduction

␤-Aspartyl and ␥-glutamyl peptides are normal constituents of mammalian urine [1, 2] and tissues [3]. The hydrolysis of ␤-aspartylglycosylamines catalyzed by glycosylasparaginase is initiated by the binding of the ␤-aspartyl moiety into the active site of the enzyme through its free ␣-amino and ␣-carboxyl groups [8, 9]. The GA-catalyzed hydrolysis of L-asparagine occurs in a similar manner, resulting in the formation of ␤-aspartyl enzyme and ammonia [12].

Results

Conclusion