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Mechanisms of protein folding and quality control in bacteria
New Biotechnology · Volume 31S · July 2014
Symposium 12: Stress responses in microbial bioprocessing O12-1 Mechanisms of protein folding and quality control in bacteria Bernd Bukau Center for Molecular Biology of the University of Heidelberg, German Cancer Research Center, DKFZ-ZMBH Alliance, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany
SYMPOSIUM 12: STRESS RESPONSES IN MICROBIAL BIOPROCESSING
References [1].Wang B, Kitney R, Joly N, Buck M. Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology. Nat Commun 2011;2:508. [2].Wang B, Buck M. Customizing cell signalling using engineered genetic logic circuits. Trends Microbiol 2012;20(8):376–84. [3].Wang B, Barahona M, Buck M. A modular cell-based biosensor using engineered genetic logic circuits to detect and integrate multiple environmental signals. Biosens Bioelectron 2013;40:368–76.
http://dx.doi.org/10.1016/j.nbt.2014.05.1714
O12-3 Protein homeostasis is established by a complex cellular machinery which assists and regulates regular folding pathways and counteracts protein misfolding and aggregation. A particularly critical process in the life of a protein is the native folding of newly synthesized proteins, which therefore is tightly controlled. Already during ongoing synthesis by the ribosome nascent polypeptides are subject to enzymatic processing, chaperone-assisted folding to the native state or targeting to translocation pores at membranes. The ribosome itself plays a key role in these different tasks by serving as platform for the regulated association of enzymes, targeting factors and chaperones that act upon the nascent polypeptides emerging from the exit tunnel. The molecular mechanisms integrating the different co-translational processes leading to the maturation and native folding of nascent chains will be described. http://dx.doi.org/10.1016/j.nbt.2014.05.1713
O12-2 Engineering customised cell signalling circuits and their biotechnological applications Baojun Wang University of Edinburgh, United Kingdom
Cells live in an ever-changing environment and continuously sense, process and react to environmental signals using their inherent signalling and gene regulatory networks. Here I will present the construction of synthetic gene circuits to customise cellular information processing and responses by harnessing the inherent modularity of signalling networks [1–3]. In particular, a set of modular and orthogonal genetic logic gates, e.g. AND and NAND, and analogue circuits such as a gain-tunable genetic amplifier were engineered to modulate multiplein vivo transcriptional signals in either digital-like or bespoke analogue manner. I will then show that how these gene circuits can be used to enhance the specificity and sensitivity of synthetic cell-based biosensors for detecting heavy metal ions and bacterial signalling molecules in an aqueous environment, and to realise robust gene expression control and sensing in single cells over a range of abiotic conditions. Furthermore, we are engineering modular genetic controllers that can act as dynamic stress sensor-regulators to achieve adaptive control of cellular pathway gene expression flows for optimised biomolecule production.
Stochastic activation of the GlpR-controlled glp gene cluster in Pseudomonas putida KT2440 results in a bistable growth pattern on glycerol Pablo Ivan Nikel ∗ , Victor de Lorenzo Centro Nacional de Biotecnologia (CNB-CSIC), Spain
Phenotypic variation is a widespread trait among prokaryotes, and the molecular mechanisms underlying the phenomenon include genetic changes (e.g., genomic inversions and strandslippage processes), epigenetic variations (e.g., distinct patterns of DNA methylation), as well as feed-back-based multi-stability. All these mechanisms are known to ultimately lead to the appearance of at least two distinct phenotypes within an otherwise isogenic population. However, this complex trait has been scarcely explored in microorganisms that have environmental and industrial interest. The soil bacterium Pseudomonas putida exhibits promising biotechnological potential, together with its generally regardedas-safe certificates, resistance to endogenous and exogenous stress, amenability to genetic manipulation, and suitability as a host for heterologous gene expression. Growth of P. putida KT2440 on glycerol is characterized by an unexpectedly long lag phase (Nikel PI, Kim J, de Lorenzo V. Metabolic and regulatory rearrangements underlying glycerol metabolism in Pseudomonas putida KT2440. Environ Microbiol 2014;16:239-54). In the present contribution, the growth of individual P. putida KT2440 cells was assessed on glycerol to further explore this phenotypic behavior. It was found that the physiological properties of cells growing on this carbon source resulted in the appearance of two distinct cell sub-populations which significantly differed in their metabolic activity. These macroscopic properties are wired to the dual logic of the GlpR regulator, repressing the transcription of the cognate glp genes, encoding the enzymes needed for glycerol catabolism. Analyzed in perspective, these results suggest that P. putida is subjected to a carbon-source-dependent bet-hedging strategy that might be relevant in the natural niches it inhabits. http://dx.doi.org/10.1016/j.nbt.2014.05.1715
www.elsevier.com/locate/nbt S45
Symposium 12: Stress responses in microbial bioprocessing O12-1 Mechanisms of protein folding and quality control in bacteria Bernd Bukau Center for Molecular Biology of the University of Heidelberg, German Cancer Research Center, DKFZ-ZMBH Alliance, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany
SYMPOSIUM 12: STRESS RESPONSES IN MICROBIAL BIOPROCESSING
References [1].Wang B, Kitney R, Joly N, Buck M. Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology. Nat Commun 2011;2:508. [2].Wang B, Buck M. Customizing cell signalling using engineered genetic logic circuits. Trends Microbiol 2012;20(8):376–84. [3].Wang B, Barahona M, Buck M. A modular cell-based biosensor using engineered genetic logic circuits to detect and integrate multiple environmental signals. Biosens Bioelectron 2013;40:368–76.
http://dx.doi.org/10.1016/j.nbt.2014.05.1714
O12-3 Protein homeostasis is established by a complex cellular machinery which assists and regulates regular folding pathways and counteracts protein misfolding and aggregation. A particularly critical process in the life of a protein is the native folding of newly synthesized proteins, which therefore is tightly controlled. Already during ongoing synthesis by the ribosome nascent polypeptides are subject to enzymatic processing, chaperone-assisted folding to the native state or targeting to translocation pores at membranes. The ribosome itself plays a key role in these different tasks by serving as platform for the regulated association of enzymes, targeting factors and chaperones that act upon the nascent polypeptides emerging from the exit tunnel. The molecular mechanisms integrating the different co-translational processes leading to the maturation and native folding of nascent chains will be described. http://dx.doi.org/10.1016/j.nbt.2014.05.1713
O12-2 Engineering customised cell signalling circuits and their biotechnological applications Baojun Wang University of Edinburgh, United Kingdom
Cells live in an ever-changing environment and continuously sense, process and react to environmental signals using their inherent signalling and gene regulatory networks. Here I will present the construction of synthetic gene circuits to customise cellular information processing and responses by harnessing the inherent modularity of signalling networks [1–3]. In particular, a set of modular and orthogonal genetic logic gates, e.g. AND and NAND, and analogue circuits such as a gain-tunable genetic amplifier were engineered to modulate multiplein vivo transcriptional signals in either digital-like or bespoke analogue manner. I will then show that how these gene circuits can be used to enhance the specificity and sensitivity of synthetic cell-based biosensors for detecting heavy metal ions and bacterial signalling molecules in an aqueous environment, and to realise robust gene expression control and sensing in single cells over a range of abiotic conditions. Furthermore, we are engineering modular genetic controllers that can act as dynamic stress sensor-regulators to achieve adaptive control of cellular pathway gene expression flows for optimised biomolecule production.
Stochastic activation of the GlpR-controlled glp gene cluster in Pseudomonas putida KT2440 results in a bistable growth pattern on glycerol Pablo Ivan Nikel ∗ , Victor de Lorenzo Centro Nacional de Biotecnologia (CNB-CSIC), Spain
Phenotypic variation is a widespread trait among prokaryotes, and the molecular mechanisms underlying the phenomenon include genetic changes (e.g., genomic inversions and strandslippage processes), epigenetic variations (e.g., distinct patterns of DNA methylation), as well as feed-back-based multi-stability. All these mechanisms are known to ultimately lead to the appearance of at least two distinct phenotypes within an otherwise isogenic population. However, this complex trait has been scarcely explored in microorganisms that have environmental and industrial interest. The soil bacterium Pseudomonas putida exhibits promising biotechnological potential, together with its generally regardedas-safe certificates, resistance to endogenous and exogenous stress, amenability to genetic manipulation, and suitability as a host for heterologous gene expression. Growth of P. putida KT2440 on glycerol is characterized by an unexpectedly long lag phase (Nikel PI, Kim J, de Lorenzo V. Metabolic and regulatory rearrangements underlying glycerol metabolism in Pseudomonas putida KT2440. Environ Microbiol 2014;16:239-54). In the present contribution, the growth of individual P. putida KT2440 cells was assessed on glycerol to further explore this phenotypic behavior. It was found that the physiological properties of cells growing on this carbon source resulted in the appearance of two distinct cell sub-populations which significantly differed in their metabolic activity. These macroscopic properties are wired to the dual logic of the GlpR regulator, repressing the transcription of the cognate glp genes, encoding the enzymes needed for glycerol catabolism. Analyzed in perspective, these results suggest that P. putida is subjected to a carbon-source-dependent bet-hedging strategy that might be relevant in the natural niches it inhabits. http://dx.doi.org/10.1016/j.nbt.2014.05.1715
www.elsevier.com/locate/nbt S45