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Expression of recombinant protease MarP from Mycobacterium tuberculosis in Pichia pastoris and its effect on human monocytes.

Authors
  • García-González, Gerardo1
  • Ascacio-Martínez, Jorge Ángel2
  • Hernández-Bello, Romel1
  • González, Gloria María1
  • Palma-Nicolás, José Prisco3
  • 1 Departamento de Microbiología, Facultad de Medicina y Hospital Universitario "Dr. José Eleuterio González", Universidad Autónoma de Nuevo León, Ave. Francisoco I. Madero y Dr. Eduardo Aguirre Pequeño s/n, Col. Mitras Centro, C.P. 64460, Monterrey, Nuevo León, Mexico. , (Mexico)
  • 2 Departamento de Bioquímica y medicina molecular, Facultad de Medicina y Hospital Universitario "Dr. José Eleuterio González", Universidad Autónoma de Nuevo León, Monterrey, Nuevo León, Mexico. , (Mexico)
  • 3 Departamento de Microbiología, Facultad de Medicina y Hospital Universitario "Dr. José Eleuterio González", Universidad Autónoma de Nuevo León, Ave. Francisoco I. Madero y Dr. Eduardo Aguirre Pequeño s/n, Col. Mitras Centro, C.P. 64460, Monterrey, Nuevo León, Mexico. [email protected] , (Mexico)
Type
Published Article
Journal
Biotechnology Letters
Publisher
Springer-Verlag
Publication Date
Sep 01, 2021
Volume
43
Issue
9
Pages
1787–1798
Identifiers
DOI: 10.1007/s10529-021-03149-3
PMID: 34028659
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

Mycobacterial acid-resistant protease (MarP) is a membrane-associated serine protease involved in the survival of Mycobacterium tuberculosis in macrophages; here we produced MarP in the yeast Pichia pastoris and study its involvement in macrophage immune modulation. Pichia pastoris vectors, harboring a full-length or a partial sequence of MarP, were constructed. GS115 clones were selected, and homologous recombination at the AOX1 locus was assessed by PCR. Protein was purified by nickel affinity chromatography, and its effect on the cytokine profile was tested in human monocytes. Only the partial MarP protein (121-397 a.a.) lacking the transmembrane domain was successfully expressed as an N-glycosylated proteolytically active protease. In vitro stimulation of THP-1 cells with MarP promoted the release of TNF-α and IL-10. Mycobacterial MarP was successfully expressed in P. pastoris, and it is capable of cytokine release in vitro. © 2021. The Author(s), under exclusive licence to Springer Nature B.V.

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