New Insights into the Determinants of Specificity in Human Type I Arginase: Generation of a Mutant That Is Only Active with Agmatine as Substrate

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dc.contributor.authorOrellana, María-Soledad
dc.contributor.authorJaña, Gonzalo A.
dc.contributor.authorFigueroa, Maximiliano
dc.contributor.authorMartínez-Oyanedel, José
dc.contributor.authorMedina, Fabiola E.
dc.contributor.authorTarifeño-Saldivia, Estefanía
dc.contributor.authorGatica, Marcell
dc.contributor.authorGarcía-Robles, María Ángeles
dc.contributor.authorCarvajal, Nelson
dc.contributor.authorUribe, Elena
dc.date.accessioned2024-04-30T15:01:42Z
dc.date.available2024-04-30T15:01:42Z
dc.date.issued2022-06-02
dc.descriptionIndexación: Scopus.
dc.description.abstractArginase catalyzes the hydrolysis of L-arginine into L-ornithine and urea. This enzyme has several analogies with agmatinase, which catalyzes the hydrolysis of agmatine into putrescine and urea. However, this contrasts with the highlighted specificity that each one presents for their respective substrate. A comparison of available crystal structures for arginases reveals an important difference in the extension of two loops located in the entrance of the active site. The first, denominated loop A (I129-L140) contains the residues that interact with the alpha carboxyl group or arginine of arginase, and the loop B (D181-P184) contains the residues that interact with the alpha amino group of arginine. In this work, to determine the importance of these loops in the specificity of arginase, single, double, and triple arginase mutants in these loops were constructed, as well as chimeras between type I human arginase and E. coli agmatinase. In previous studies, the substitution of N130D in arginase (in loop A) generated a species capable of hydrolyzing arginine and agmatine. Now, the specificity of arginase is completely altered, generating a chimeric species that is only active with agmatine as a substrate, by substituting I129T, N130Y, and T131A together with the elimination of residues P132, L133, and T134. In addition, Quantum Mechanic/Molecular Mechanic (QM/MM) calculations were carried out to study the accommodation of the substrates in in the active site of this chimera. With these results it is concluded that this loop is decisive to discriminate the type of substrate susceptible to be hydrolyzed by arginase. Evidence was also obtained to define the loop B as a structural determinant for substrate affinity. Concretely, the double mutation D181T and V182E generate an enzyme with an essentially unaltered kcat value, but with a significantly increased Km value for arginine and a significant decrease in affinity for its product ornithine. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.
dc.description.urihttps://www.mdpi.com/1422-0067/23/12/6438
dc.identifier.citationInternational Journal of Molecular Sciences, Volume 23, Issue 12, June-2 2022, Article number 6438
dc.identifier.doi10.3390/ijms23126438
dc.identifier.issn1661-6596
dc.identifier.urihttps://repositorio.unab.cl/handle/ria/56483
dc.language.isoen
dc.publisherMDPI
dc.rights.licenseCC BY 4.0 DEED Atribución 4.0 Internacional
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/deed.es
dc.subjectAgmatine
dc.subjectArginase
dc.subjectArginine
dc.subjectDeterminants of specificity
dc.titleNew Insights into the Determinants of Specificity in Human Type I Arginase: Generation of a Mutant That Is Only Active with Agmatine as Substrate
dc.typeArtículo
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