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Functional identification of a Streptomyces lividans FKBP-like protein involved in the folding of overproduced secreted proteins.

ABSTRACT: Some bacterial peptidyl-prolyl cis/trans isomerases (PPIases) are involved in secretory protein folding after the translocation step. Streptomyces lividans has been used as a host for engineering extracellular overproduction of homologous and heterologous proteins in industrial applications. Although the mechanisms governing the major secretory pathway (Sec route) and the minor secretory pathway (Tat route) are reasonably well described, the function of proteins responsible for the extracellular secretory protein folding is not characterized as yet. We have characterized a Tat-dependent S. lividans FK506-binding protein-like lipoprotein (FKBP) that has PPIase activity. A mutant in the sli-fkbp gene induces a secretion stress response and affects secretion and activity of the Sec-dependent protein ?-amylase. Additionally, propagation in high copy number of the sli-fkbp gene has a positive effect on the activity of both the overproduced ?-amylase and the overproduced Tat-dependent agarase, both containing proline cis isomers. Targeted proteomic analyses showed that a relevant group of secreted proteins in S. lividans TK21 are affected by Sli-FKBP, revealing a wide substrate range. The results obtained indicate that, regardless of the secretory route used by proteins in S. lividans, adjusting the expression of sli-fkbp may facilitate folding of dependent proteins when engineering Streptomyces strains for the overproduction of homologous or heterologous secretory proteins.

SUBMITTER: Vicente RL

PROVIDER: S-EPMC6833217 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Functional identification of a Streptomyces lividans FKBP-like protein involved in the folding of overproduced secreted proteins.

Vicente R L RL Marín S S Valverde J R JR Palomino C C Mellado R P RP Gullón S S

Open biology 20191030 10

Some bacterial peptidyl-prolyl cis/trans isomerases (PPIases) are involved in secretory protein folding after the translocation step. Streptomyces lividans has been used as a host for engineering extracellular overproduction of homologous and heterologous proteins in industrial applications. Although the mechanisms governing the major secretory pathway (Sec route) and the minor secretory pathway (Tat route) are reasonably well described, the function of proteins responsible for the ...[more]

PMID: 31662098

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Project description:BACKGROUND:Streptomyces lividans is an appealing host for the production of proteins of biotechnological interest due to its relaxed exogenous DNA restriction system and its ability to secrete proteins directly to the medium through the major Sec or the minor Tat routes. Often, protein secretion displays non-uniform time-dependent patterns. Understanding the associated metabolic changes is a crucial step to engineer protein production. Dynamic Flux Balance Analysis (DFBA) allows the study of the interactions between a modelled organism and its environment over time. Existing methods allow the specification of initial model and environment conditions, but do not allow introducing arbitrary modifications in the course of the simulation. Living organisms, however, display unexpected adaptive metabolic behaviours in response to unpredictable changes in their environment. Engineering the secretion of products of biotechnological interest has systematically proven especially difficult to model using DFBA. Accurate time-dependent modelling of complex and/or arbitrary, adaptive metabolic processes demands an extended approach to DFBA. RESULTS:In this work, we introduce Adaptive DFBA, a novel, versatile simulation approach that permits inclusion of changes in the organism or the environment at any time in the simulation, either arbitrary or interactively responsive to environmental changes. This approach extends traditional DFBA to allow steering arbitrarily complex simulations of metabolic dynamics. When applied to Sec- or Tat-dependent secretion of overproduced proteins in S. lividans, Adaptive DFBA can overcome the limitations of traditional DFBA to reproduce experimental data on plasmid-free, plasmid bearing and secretory protein overproducing S. lividans TK24, and can yield useful insights on the behaviour of systems with limited experimental knowledge such as agarase or amylase overproduction in S. lividans TK21. CONCLUSIONS:Adaptive DFBA has allowed us to overcome DFBA limitations and to generate more accurate models of the metabolism during the overproduction of secretory proteins in S. lividans, improving our understanding of the underlying processes. Adaptive DFBA is versatile enough to permit dynamical metabolic simulations of arbitrarily complex biotechnological processes.

Dataset Information

Functional identification of a Streptomyces lividans FKBP-like protein involved in the folding of overproduced secreted proteins.

Publications

Functional identification of a <i>Streptomyces lividans</i> FKBP-like protein involved in the folding of overproduced secreted proteins.

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