Bacterial luciferase alpha beta fusion protein is fully active as a monomer and highly sensitive in vivo to elevated temperature.

Escher, Alan; O’Kane, Dennis J.; Lee, John; Szalay, Aladar A.

doi:10.1073/pnas.86.17.6528

Cited by 81 publications

(50 citation statements)

References 21 publications

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“…Bacterial luciferase luxAB gene, derived from Vibrio harveyi luxA and luxB, was PCR-cloned from pDCM58, a kind gift from Dr. Daniel C. Masison (National Institutes of Health, Bethesda, MD) (50,51).…”

Section: Methodsmentioning

confidence: 99%

Neuronal Calcium Sensor-1 (Ncs1p) Is Up-regulated by Calcineurin to Promote Ca2+ Tolerance in Fission Yeast

Hamasaki-Katagiri

Ames

2010

Journal of Biological Chemistry

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) regulates signaling processes in the fission yeast, Schizosaccharomyces pombe (reviewed in Refs. 1-3), as it does in mammalian cells (reviewed in Ref. 4). Fission yeast contains ion channels (5, 6) and transporters (7-9) homologous to those involved in regulating Ca 2ϩ entry and signal transduction in excitable cells. Thus, stimulation of yeast cells by pheromones or other extracellular stimuli lead to an increase in intracellular Ca 2ϩ levels (10, 11) sensed by Ca 2ϩ -binding proteins belonging to the EF-hand superfamily (12). In S. pombe, a total of 23 genes specifies EF-hand-containing Ca 2ϩ -binding proteins, including cam1 (calmodulin) (13), cnb1 (calcineurin B) (14), and the recently characterized ncs1 gene (Ncs1p) (15), belonging to the neuronal calcium sensor (NCS) 2 family (16) expressed mostly in the central nervous system (17, 18).The primary sequence of S. pombe Ncs1p is Ͼ50% identical to neuronal homologs of the NCS branch of the EF-hand superfamily (19,20). Recoverin, the most intensively studied NCS protein, serves as a Ca 2ϩ sensor in retinal rod and cone cells where it controls the desensitization of rhodopsin (21-24). The NCS family also includes neuronal Ca 2ϩ sensors such as neurocalcin (25), hippocalcin (26), and frequenin (27), as well as the budding yeast homolog, Frq1 (28). All members of the NCS family have around 200 amino acid residues, contain N-terminal myristoylation, and possess four EF-hands.Three-dimensional structures are now known for many NCS proteins, including recoverin (29, 30), frequenin (31), Frq1 (32), neurocalcin (33), and guanylate cyclase activator proteins (34, 35). The Ca 2ϩ -bound forms of these proteins all share a common fold with four EF-hands arranged in a tandem array, with an exposed N-terminal myristoyl group. Binding of Ca 2ϩ to recoverin leads to extrusion of its myristoyl group such that many hydrophobic residues become exposed for target recognition (30, 36). The Ca 2ϩ -induced exposure of the myristoyl group, termed the Ca 2ϩ -myristoyl switch, enables recoverin to bind to membrane targets only at high Ca 2ϩ levels (37, 38). NCS proteins interact with and regulate many different physiological targets. Recoverin binds specifically and regulates rhodopsin kinase (23), whereas the guanylate cyclase activator proteins specifically regulate retinal guanylate cyclases (39, 40). NCS-1 and yeast Frq1 activate phosphatidylinositol 4-kinase important for secretion from the Golgi (28). Hippocalcin and VILIP proteins regulate guanylate cyclase and nicotinamide acetylcholine receptors implicated in synaptic plasticity (41). KChIPs (42), hippocalcin (43), and NCS-1 (44) all bind and regulate various ion channels and thus serve as Ca 2ϩ sensors that control membrane excitability. The mechanism of target regulation in each case is not understood, and it remains unclear how the structurally similar NCS proteins are able to specifically bind and regulate so many different target proteins.Yeast genetics is a powerful tool for dissecting NCS protein interactions in signa...

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Section: Methodsmentioning

confidence: 99%

Neuronal Calcium Sensor-1 (Ncs1p) Is Up-regulated by Calcineurin to Promote Ca2+ Tolerance in Fission Yeast

Hamasaki-Katagiri

Ames

2010

Journal of Biological Chemistry

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“…When the growth temperature was decreased to a tolerable, but not ideal temperature of 30°C, bioluminescent detection increased 10-fold (Pazzagli et al, 1992). The temperaturedependent nature of this bioluminescent decrease was additionally confirmed through further testing in E. coli, where it was determined that hosts expressing LuxA-LuxB fusion proteins were capable of producing a greater than 50,000-fold increase in bioluminescent production when grown at 23°C compared to growth at 37°C (Escher et al, 1989). This highlights the need to not only evaluate the potential genetic hurdles to exogenous expression of a target gene, but also to consider the physiological limitations constraining expression of the protein encoded from that gene as well.…”

Section: Initial Attempts At Exogenous Expressionmentioning

confidence: 79%

Expression of Non-Native Genes in a Surrogate Host Organism

Close¹,

Xu²,

Smartt³

et al. 2012

Genetic Engineering - Basics, New Applications and Responsibilities

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“…Multimerization is likely to arise from the need for functionality, and such a need varies significantly between proteins. Some proteins are functional both in monomeric as well as in various multimer forms [7,8], while others are only functional in a specific form [9]. …”

Section: Introductionmentioning

confidence: 99%

Effects of multimerization on the temporal variability of protein complex abundance

et al. 2013

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We explore whether the process of multimerization can be used as a means to regulate noise in the abundance of functional protein complexes. Additionally, we analyze how this process affects the mean level of these functional units, response time of a gene, and temporal correlation between the numbers of expressed proteins and of the functional multimers. We show that, although multimerization increases noise by reducing the mean number of functional complexes it can reduce noise in comparison with a monomer, when abundance of the functional proteins are comparable. Alternatively, reduction in noise occurs if both monomeric and multimeric forms of the protein are functional. Moreover, we find that multimerization either increases the response time to external signals or decreases the correlation between number of functional complexes and protein production kinetics. Finally, we show that the results are in agreement with recent genome-wide assessments of cell-to-cell variability in protein numbers and of multimerization in essential and non-essential genes in Escherichia coli, and that the effects of multimerization are tangible at the level of genetic circuits.

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Bacterial luciferase alpha beta fusion protein is fully active as a monomer and highly sensitive in vivo to elevated temperature.

Abstract: Bacterial luciferase a13 fusion protein is fully active as a monomer and highly sensitive in vivo to elevated temperature (

Cited by 81 publications

References 21 publications

Neuronal Calcium Sensor-1 (Ncs1p) Is Up-regulated by Calcineurin to Promote Ca2+ Tolerance in Fission Yeast

Neuronal Calcium Sensor-1 (Ncs1p) Is Up-regulated by Calcineurin to Promote Ca2+ Tolerance in Fission Yeast

Expression of Non-Native Genes in a Surrogate Host Organism

Effects of multimerization on the temporal variability of protein complex abundance

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