Detection of Single Oxygen Molecules with Fluorescence-Labeled Hemocyanins

Erker, Wolfgang; Sdorra, Sven; Basché, Thomas

doi:10.1021/ja053720k

Cited by 20 publications

(11 citation statements)

References 15 publications

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“…We were able to image individual but low resolution molecules of this hemocyanin after immobilization in an almost native state in air. Functionality of adsorbed 4 Â 6-mer hemocyanins has recently been shown by single-molecule fluorescence microscopy (Erker et al, 2005b). Oxygen binding of single hemocyanin molecules, imaged at a glass-water interface was demonstrated.…”

Section: Resultsmentioning

confidence: 97%

Immobilization and AFM of single 4×6-mer tarantula hemocyanin molecules

Erker¹,

Scheumann

Möller³

et al. 2006

Micron

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Section: Resultsmentioning

confidence: 97%

Immobilization and AFM of single 4×6-mer tarantula hemocyanin molecules

Erker¹,

Scheumann

Möller³

et al. 2006

Micron

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“…Erker, Basché, and coworkers used this strategy to study oxygen binding by binuclear copper protein hemocyanin (108, 109). Aartsman, Canters, Schmidt and coworkers used this strategy to probe the redox states of blue copper protein azurin (110, 111).…”

Section: Related Single-molecule Bioinorganic Workmentioning

confidence: 99%

Single-Molecule Dynamics and Mechanisms of Metalloregulators and Metallochaperones

et al. 2013

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Understanding how cells regulate and transport metal ions is an important goal in the field of bioinorganic chemistry, a frontier research area that resides at the interfaces of chemistry and biology. This Current Topics article reviews recent advances from the authors' group in using single-molecule fluorescence imaging techniques to identify the mechanisms of metal homeostatic proteins, including metalloregulators and metallochaperones. It emphasizes the novel mechanistic insights into how dynamic protein–DNA and protein–protein interactions offer efficient pathways for MerR-family metalloregulators and copper chaperones to fulfill their functions. The article also summarizes other related single-molecule studies of bioinorganic systems, and gives an outlook toward single-molecule imaging of metalloprotein functions in living cells.

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“…All measurements were made in PBS buffer under ambient conditions using two different confocal fluorescence microscopes. On the one hand, a custom-built microscope (based on a Zeiss Axiovert 135 TV inverted microscope) was used, which has single-molecule sensitivity (Erker et al, 2005;Kulzer et al, 1999). Images with a typical size of 10 mm610 mm and 1286128 pixels were recorded with an integration time of 5 ms per pixel (objective: Zeiss Plan-Neofluar 1006/1.30 oil).…”

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confidence: 99%

Polar accumulation of the metabolic sensory histidine kinases DcuS and CitA in Escherichia coli

et al. 2008

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Signal transduction in prokaryotes is frequently accomplished by two-component regulatory systems in which a histidine protein kinase is the sensory component. Many of these sensory kinases control metabolic processes that do not show an obvious requirement for inhomogeneous distribution within bacterial cells. Here, the sensory kinases DcuS and CitA, two histidine kinases of Escherichia coli, were investigated. Both are membrane-integral and involved in the regulation of carboxylate metabolism. The two-component sensors were fused with yellow fluorescent protein (YFP) and live images of immobilized cells were obtained by confocal laser fluorescence microscopy. The fluorescence of the fusion proteins was concentrated at the poles of the cells, indicating polar accumulation of the sensory kinases. For quantitative evaluation, line profiles of the imaged fluorescence intensities were generated; these revealed that the fluorescence intensity of the polar bright spots was 2.3-8.5 times higher than that of the cytoplasm. With respect to the cylindrical part of the membrane, the values were lower by about 40 %. The polar accumulation was comparable to that of methyl-accepting chemotaxis proteins (MCPs) and MCP-related proteins. The degree of DcuS-YFP localization was independent of the presence of MCP and the expression level of dcuS-yfp (or DcuS concentration). The presence of effector (fumarate or citrate, respectively) increased the polar accumulation by more than 20 %. Cell fractionation demonstrated that polar accumulation was not related to inclusion body formation. Therefore, sensory kinases DcuS and CitA, which regulate metabolic processes without obvious polar function, exhibit polar accumulation.

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Detection of Single Oxygen Molecules with Fluorescence-Labeled Hemocyanins

Cited by 20 publications

References 15 publications

Immobilization and AFM of single 4×6-mer tarantula hemocyanin molecules

Immobilization and AFM of single 4×6-mer tarantula hemocyanin molecules

Single-Molecule Dynamics and Mechanisms of Metalloregulators and Metallochaperones

Polar accumulation of the metabolic sensory histidine kinases DcuS and CitA in Escherichia coli

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