Probing heat and oxidation induced conformational changes of molecular chaperone artemin by excitation-emission fluorescence spectroscopy

Takalloo, Zeinab; Masroor, Mohammad Javad; Mani-Varnosfaderani, Ahmad; Maroufi, Bahman; Sajedi, Reza H.

doi:10.1016/j.jphotobiol.2020.112013

Cited by 5 publications

(10 citation statements)

References 43 publications

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“…The detection of peak T and/or peak B provided an indication of different protein structures due to fundamental amino acid variability. 62,63 FEEM could thus indicate potential differences among the proteins by identifying different fluorescent amino acids present using the peak-picking method.…”

Section: Resultsmentioning

confidence: 99%

Considerations of the limitations of commonly applied characterisation methods in understanding protein-driven irreversible fouling

Mustafa

Rao

Henderson

et al. 2022

Environ. Sci.: Water Res. Technol.

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Section: Resultsmentioning

confidence: 99%

Considerations of the limitations of commonly applied characterisation methods in understanding protein-driven irreversible fouling

Mustafa

Rao

Henderson

et al. 2022

Environ. Sci.: Water Res. Technol.

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“…Movies were collected at 130,000× magnification in super resolution mode resulting in a pixel size of 0.3398 Å respectively. Movies were collected at a total dose ranging from 41.7 to 58.9 e - /Å 2 , with 0.5 to 1.8 s exposures, and a defocus range between - 0.3 to -1.3 µm.…”

Section: Methodsmentioning

confidence: 99%

“…The key to surviving such harsh conditions has been tracked to the brine shrimp’s ability as a cyst to enter a state of metabolic hypoactivity called diapause. In this state, the cyst can survive desiccation, high and low temperatures, radiation and years of anoxia 2 . A complement of stress tolerance proteins have been reported in Artemia during diapause including p26, artemin and hsc70 1 .…”

Section: Introductionmentioning

confidence: 99%

Cryo-EM structure of the diapause chaperone artemin

Parvate

Powell

Brookreson

et al. 2022

Preprint

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The protein artemin constitutes over 10% of all protein in Artemia cysts during diapause and acts as both an RNA and protein chaperone. However, its mechanistic details remain elusive since no high-resolution structure of artemin exists. Here we report the full-length structure of artemin at 2.04 Å resolution. The cryo-EM map contains density for an intramolecular disulfide bond between Cys22-Cys61 and resolves the entire C-terminus extending into the core of the assembled protein cage. We also provide data supporting the role of C-terminal helix F towards stabilizing the dimer form that is believed to be important for its chaperoning activity. We were able to destabilize this effect by placing a tag at the C-terminus to fully pack the internal cavity and cause limited steric hindrance.

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“…Além da análise em termos de estrutura terciária global, o ambiente terciário local das proteínas HspA5 e HspA8 também foi estudado por meio da técnica de emissão de fluorescência intrínseca (TAKALLOO et al, 2020) Fonte: Autoria própria.…”

Section: Caracterização Da Estrutura Secundária E Terciáriaunclassified

“…estrutura terciária local das proteínas recombinantes HspA5 e HspA8 foi monitorada por emissão de fluorescência intrínseca do triptofano(TAKALLOO et al, 2020), utilizando um espectrofotômetro de fluorescência F-4500 (Hitachi). As amostras proteicas (3 μM), na presença ou ausência de 8 M de ureia, foram acondicionadas em uma cubeta de quartzo 1x0,2 cm e excitadas em comprimento de onda de 295 nm.Ensaios na presença dos nucleotídeos adenosina também foram realizados por fluorescência, usando as mesmas condições descritas na seção anterior.…”

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Caracterização estrutural, funcional e estabilidade térmica das proteínas humanas HspA5 e HspA8: uma abordagem comparativa

Silva¹

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Heat shock proteins (Hsps) are important components of cell protein quality control (PQC), acting on protein synthesis, folding, traffic and degradation. The central component in the PQC is the Hsp70 family, which consists of highly conserved, ubiquitous and ATP-dependent molecular chaperones. These chaperones can assume different conformations that are regulated by their dynamic association with the adenosine nucleotides (ATP and ADP), as well as by their interaction with cochaperones and client proteins. In humans there are several Hsp70 proteins, most of which are found in the cytoplasm, while others are specific for organelles. The objective of this work was to structurally and functionally characterize the cytosolic (HspA8) and endoplasmic reticulum (HspA5) human Hsp70 proteins, using similar methodologies.For this, the recombinant hHsp70 proteins were expressed, purified and then characterized for structure, stability and function by means of biophysical tests. The results show that the HspA5 and HspA8 proteins were produced in their folded form, with secondary structure content rich in α-helices and as slightly elongated monomers in solution. Such Hsp70 showed similar chemical and thermal stability, however two thermal transitions (Tm) were observed for HspA5 and three for HspA8, indicating structural differences that result in distinct thermal stability. Changes in secondary and tertiary structure were observed in the proteins under study in the presence of nucleotides, which also caused the thermal stabilization of hHsp70. The presence of Ca 2+ and Mg 2+ ions favored the binding of hHsp70 to ATP and ADP nucleotides in different ways, and unlike HspA5, the HspA8 protein interacted with Ca 2+ even in the absence of nucleotides. A process thermodynamically governed by enthalpy and entropy was observed for the interaction of the adenosine nucleotides with the HspA5 and HspA8 proteins. hHsp70 showed similar and relatively low ATP activity, however the affinity of HspA5 for ATP was higher than that of HspA8. Additionally, the HspA5 and HspA8 proteins were effective in preventing the aggregation of model client proteins, with different specificities. In summary, the data obtained demonstrate that, although they share 66% of identity, the human proteins HspA5 and HspA8 present structural and functional peculiarities, thus enriching the knowledge about such macromolecules.

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Probing heat and oxidation induced conformational changes of molecular chaperone artemin by excitation-emission fluorescence spectroscopy

Cited by 5 publications

References 43 publications

Considerations of the limitations of commonly applied characterisation methods in understanding protein-driven irreversible fouling

Considerations of the limitations of commonly applied characterisation methods in understanding protein-driven irreversible fouling

Cryo-EM structure of the diapause chaperone artemin

Caracterização estrutural, funcional e estabilidade térmica das proteínas humanas HspA5 e HspA8: uma abordagem comparativa

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