The role of ATP hydrolysis in the function of the chaperonin GroEL: dynamic complex formation with GroES

1. The intrinsic Na+, K+, Mg2+ and Ca2+ contents of a preparation of membrane fragments from ox brain were determined by emission flame photometry. 2. Centrifugal washing of the preparation with imidazole-buffered EDTA solutions decreased the bound Na+ from 90±20 to 24±12, the bound K+ from 27±3 to 7±2, the bound Mg2+ from 20±2 to 3±1 and the bound calcium from 8±1 to <1nmol/mg of protein. 3. The activities of the Na++K++Mg2+-stimulated adenosine triphosphatase and the Na+-dependent reaction forming bound phosphate were compared in the unwashed and washed preparations at an ATP concentration of 2.5μm (ATP/protein ratio 12.5pmol/μg). 4. The Na+-dependent hydrolysis of ATP as well as the plateau concentration of bound phosphate and the rate of dephosphorylation were decreased in the washed preparation. The time-course of formation and decline of bound phosphate was fully restored by the addition of 2.5μm-magnesium chloride and 2μm-potassium chloride. Addition of 2.5μm-magnesium chloride alone fully restored the plateau concentration of bound phosphate, but the rate of dephosphorylation was only slightly increased. Na+-dependent ATP hydrolysis was partly restored with 2.5μm-magnesium chloride; addition of K+ in the range 2–10μm-potassium chloride then further restored hydrolysis but not to the control rate. 5. Pretreatment of the washed preparation at 0°C with 0.5nmol of K+/mg of protein so that the final added K+ in the reaction mixture was 0.1μm restored the Na+-dependent hydrolysis of ATP and the time-course of the reaction forming bound phosphate. 6. The binding of [42K]potassium chloride by the washed membrane preparation was examined. Binding in a solution containing 10nmol of K+/mg of protein was linear over a period of 20min and was inhibited by Na+. Half-maximal inhibition of 42K+-binding required a 100-fold excess of sodium chloride. 7. It was concluded (a) that a significant fraction of the apparent Na+-dependent hydrolysis of ATP observed in the unwashed preparation is due to activation by bound K+ and Mg2+ of the Na++K++Mg2+-stimulated adenosine triphosphatase system and (b) that the enzyme system is able to bind K+ from a solution of 0.5μm-potassium chloride.

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Impact of Quantum Dot Surface on Complex Formation with Chlorin e6 and Photodynamic Therapy

Nanomaterials ◽

10.3390/nano9010009 ◽

2018 ◽

Vol 9 (1) ◽

pp. 9 ◽

Cited By ~ 2

Author(s):

Artiom Skripka ◽

Dominyka Dapkute ◽

Jurga Valanciunaite ◽

Vitalijus Karabanovas ◽

Ricardas Rotomskis

Keyword(s):

Photodynamic Therapy ◽

Cancer Therapy ◽

Complex Formation ◽

Chemical Properties ◽

Disease Diagnosis ◽

Chlorin E6 ◽

Deep Tissue ◽

Physico Chemical

Nanomaterials have permeated various fields of scientific research, including that of biomedicine, as alternatives for disease diagnosis and therapy. Among different structures, quantum dots (QDs) have distinctive physico-chemical properties sought after in cancer research and eradication. Within the context of cancer therapy, QDs serve the role of transporters and energy donors to photodynamic therapy (PDT) drugs, extending the applicability and efficiency of classic PDT. In contrast to conventional PDT agents, QDs’ surface can be designed to promote cellular targeting and internalization, while their spectral properties enable better light harvesting and deep-tissue use. Here, we investigate the possibility of complex formation between different amphiphilic coating bearing QDs and photosensitizer chlorin e6 (Ce6). We show that complex formation dynamics are dependent on the type of coating—phospholipids or amphiphilic polymers—as well as on the surface charge of QDs. Förster’s resonant energy transfer occurred in every complex studied, confirming the possibility of indirect Ce6 excitation. Nonetheless, in vitro PDT activity was restricted only to negative charge bearing QD-Ce6 complexes, correlating with better accumulation in cancer cells. Overall, these findings help to better design such and similar complexes, as gained insights can be straightforwardly translated to other types of nanostructures—expanding the palette of possible therapeutic agents for cancer therapy.

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The role of Glu259 in Escherichia coli elongation factor Tu in ternary complex formation

Protein Engineering Design and Selection ◽

10.1093/protein/11.2.101 ◽

1998 ◽

Vol 11 (2) ◽

pp. 101-108 ◽

Cited By ~ 7

Author(s):

G. N. Pedersen ◽

T. Rattenborg ◽

C. R. Knudsen ◽

B. F. Clark

Keyword(s):

Escherichia Coli ◽

Complex Formation ◽

Ternary Complex ◽

Elongation Factor ◽

Elongation Factor Tu ◽

Ternary Complex Formation

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Iron Requirement for Mn(II) Oxidation by Leptothrix discophora SS-1

Applied and Environmental Microbiology ◽

10.1128/aem.02291-08 ◽

2008 ◽

Vol 75 (5) ◽

pp. 1229-1235 ◽

Cited By ~ 28

Author(s):

Iman A. El Gheriany ◽

Daniela Bocioaga ◽

Anthony G. Hay ◽

William C. Ghiorse ◽

Michael L. Shuler ◽

...

Keyword(s):

Complex Formation ◽

Temporal Variations ◽

Iron Limitation ◽

Gene Transcript ◽

Cellular Functions ◽

Transcript Levels ◽

Leptothrix Discophora ◽

Geochemical Cycling ◽

Iron Requirement

ABSTRACT A common form of biocatalysis of Mn(II) oxidation results in the formation of biogenic Mn(III, IV) oxides and is a key reaction in the geochemical cycling of Mn. In this study, we grew the model Mn(II)-oxidizing bacterium Leptothrix discophora SS-1 in media with limited iron (0.1 μM iron/5.8 mM pyruvate) and sufficient iron (0.2 μM iron/5.8 mM pyruvate). The influence of iron on the rate of extracellular Mn(II) oxidation was evaluated. Cultures in which cell growth was limited by iron exhibited reduced abilities to oxidize Mn(II) compared to cultures in medium with sufficient iron. While the extracellular Mn(II)-oxidizing factor (MOF) is thought to be a putative multicopper oxidase, Mn(II) oxidation in the presence of zero added Cu(II) was detected and the decrease in the observed Mn(II) oxidation rate in iron-limited cultures was not relieved when the medium was supplemented with Cu(II). The decline of Mn(II) oxidation under iron-limited conditions was not accompanied by siderophore production and is unlikely to be an artifact of siderophore complex formation with Mn(III). The temporal variations in mofA gene transcript levels under conditions of limited and abundant iron were similar, indicating that iron limitation did not interfere with the transcription of the mofA gene. Our quantitative PCR results provide a step forward in understanding the regulation of Mn(II) oxidation. The mechanistic role of iron in Mn(II) oxidation is uncertain; the data are consistent with a direct requirement for iron as a component of the MOF or an indirect effect of iron resulting from the limitation of one of many cellular functions requiring iron.

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