Controlled production of stable heterologous proteins in Lactococcus lactis

A Miyoshi; I Poquet; V Azevedo; J Commissaire; L Bermudez-Humaran; E Domakova; Y Le Loir; S C Oliveira; A Gruss; P Langella

doi:10.1128/AEM.68.6.3141-3146.2002

Controlled production of stable heterologous proteins in Lactococcus lactis

Appl Environ Microbiol. 2002 Jun;68(6):3141-6. doi: 10.1128/AEM.68.6.3141-3146.2002.

Authors

A Miyoshi¹, I Poquet, V Azevedo, J Commissaire, L Bermudez-Humaran, E Domakova, Y Le Loir, S C Oliveira, A Gruss, P Langella

Affiliation

¹ Unité de Recherches Laitières et de Génétique Appliquée, INRA, Domaine de Vilvert, 78352 Jouy en Josas cedex, France.

Abstract

The use of Lactococcus lactis (the most extensively characterized lactic acid bacterium) as a delivery organism for heterologous proteins is, in some cases, limited by low production levels and poor-quality products due to surface proteolysis. In this study, we combined in one L. lactis strain use of the nisin-inducible promoter P(nisA) and inactivation of the extracellular housekeeping protease HtrA. The ability of the mutant strain, designated htrA-NZ9000, to produce high levels of stable proteins was confirmed by using the staphylococcal nuclease (Nuc) and the following four heterologous proteins fused or not fused to Nuc that were initially unstable in wild-type L. lactis strains: (i) Staphylococcus hyicus lipase, (ii) the bovine rotavirus antigen nonstructural protein 4, (iii) human papillomavirus antigen E7, and (iv) Brucella abortus antigen L7/L12. In all cases, protein degradation was significantly lower in strain htrA-NZ9000, demonstrating the usefulness of this strain for stable heterologous protein production.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Antigens, Bacterial / genetics
Antigens, Bacterial / metabolism
Bacterial Proteins / metabolism*
Brucella abortus / chemistry
DNA-Directed RNA Polymerases*
Endonucleases / genetics
Endonucleases / metabolism
Heat-Shock Proteins*
Lactococcus lactis / genetics
Lactococcus lactis / metabolism*
Micrococcal Nuclease*
Oncogene Proteins, Viral / genetics
Oncogene Proteins, Viral / metabolism
Papillomaviridae / genetics
Periplasmic Proteins*
Recombinant Proteins / metabolism
Rotavirus / metabolism
Serine Endopeptidases / genetics
Serine Endopeptidases / metabolism
Viral Nonstructural Proteins / genetics
Viral Nonstructural Proteins / metabolism

Substances

Antigens, Bacterial
Bacterial Proteins
E7 protein, Human papillomavirus type 33
Heat-Shock Proteins
Oncogene Proteins, Viral
Periplasmic Proteins
Recombinant Proteins
Viral Nonstructural Proteins
nuc protein, staphylococcus
DNA-Directed RNA Polymerases
Endonucleases
Micrococcal Nuclease
DegP protease
Serine Endopeptidases