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The differential ability of asparagine and glutamine in promoting the closed/active enzyme conformation rationalizes the Wolinella succinogenes L-asparaginase substrate specificity.


ABSTRACT: Many side effects of current FDA-approved L-asparaginases have been related to their secondary L-glutaminase activity. The Wolinella succinogenes L-asparaginase (WoA) has been reported to be L-glutaminase free, suggesting it would have fewer side effects. Unexpectedly, the WoA variant with a proline at position 121 (WoA-P121) was found to have L-glutaminase activity in contrast to Uniprot entry P50286 (WoA-S121) that has a serine residue at this position. Towards understanding how this residue impacts the L-glutaminase property, kinetic analysis was coupled with crystal structure determination of these WoA variants. WoA-S121 was confirmed to have much lower L-glutaminase activity than WoA-P121, yet both showed comparable L-asparaginase activity. Structures of the WoA variants in complex with L-aspartic acid versus L-glutamic acid provide insights into their differential substrate selectivity. Structural analysis suggests a mechanism by which residue 121 impacts the conformation of the conserved tyrosine 27, a component of the catalytically-important flexible N-terminal loop. Surprisingly, we could fully model this loop in either its open or closed conformations, revealing the roles of specific residues of an evolutionary conserved motif among this L-asparaginase family. Together, this work showcases critical residues that influence the ability of the flexible N-terminal loop for adopting its active conformation, thereby effecting substrate specificity.

SUBMITTER: Nguyen HA 

PROVIDER: S-EPMC5282591 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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The differential ability of asparagine and glutamine in promoting the closed/active enzyme conformation rationalizes the Wolinella succinogenes L-asparaginase substrate specificity.

Nguyen Hien Anh HA   Durden Donald L DL   Lavie Arnon A  

Scientific reports 20170131


Many side effects of current FDA-approved L-asparaginases have been related to their secondary L-glutaminase activity. The Wolinella succinogenes L-asparaginase (WoA) has been reported to be L-glutaminase free, suggesting it would have fewer side effects. Unexpectedly, the WoA variant with a proline at position 121 (WoA-P<sub>121</sub>) was found to have L-glutaminase activity in contrast to Uniprot entry P50286 (WoA-S<sub>121</sub>) that has a serine residue at this position. Towards understand  ...[more]

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