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Spectroscopic and thermodynamic properties of recombinant heat shock protein A6 from Camelus dromedarius

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Abstract

Heat shock protein A6, also known as HSP70B’, is a member of the Hsp70 family of molecular chaperones. Under stressed conditions, the level of HSPA6 increases substantially, and the protein has been targeted as a biomarker of cellular stress in several studies. We report the spectroscopic and thermodynamic properties of Arabian camel species cHSPA6, determined by measurement of intrinsic and extrinsic fluorescence emission, and use of far-UV circular dichroism and dynamic multimode spectroscopy. Our results showed that cHSPA6 has similar binding affinity for both ATP and ADP (K D = ~50 nM). Binding of ATP and ADP reduced the surface hydrophobicity of the protein, and slightly altered its secondary structure, suggesting localized conformational rearrangement after ATP or ADP binding. Dynamic multimode spectroscopy revealed that cHSPA6 unfolds through three transitions with melting points (T m) of 42.3 ± 0.2, 61.3 ± 0.1, and 81.2 ± 0.2 °C. To the best of the author’s knowledge, and literature search, this is the first report of the spectroscopic and thermodynamic properties of the Arabian camel heat shock protein.

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Abbreviations

Amp:

Ampicillin

DTT:

Dithiothreitol

EDTA:

Ethylenediaminetetraacetic acid

Em:

Emission

Ex:

Excitation

FPLC:

Fast protein liquid chromatography

GdnHCl:

Guanidinium hydrochloride

IPTG:

Isopropyl β-D-1-thiogalactopyranoside

K D :

Dissociation constant

LB:

Luria–Bertani

2XLB:

Double strength Luria–Bertani

NB:

Nutrient broth

MWCO:

Molecular weight cut off

Ni-NTA:

Nickel-nitrilotriacetic acid

NBD:

Nucleotide binding domain

OD600 :

Optical density at 600 nm

PMSF:

Phenylmethylsulfonyl fluoride

rpm:

Rotation per minute

SBD:

Substrate binding domain

TB:

Terrific broth

T m :

Melting temperature

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Acknowledgments

The authors extend their appreciation to King Abdulaziz City for Science and Technology (KACST), Riyadh, Saudi Arabia, for funding this work through project # A-C-11-0606.

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Authors and Affiliations

  1. Protein Research Chair, Department of Biochemistry, College of Science, King Saud University, PO Box 2455, Riyadh, 11451, Saudi Arabia

    Ajamaluddin Malik, Haseeb Jagirdar, Abdulrahman M. Alsenaidy & Mohammad D. Bazzi

  2. Tabuk Pharmaceuticals Manufacturing Co., P.O. Box 4640, 2nd Industrial City, Dammam, 31421, Saudi Arabia

    Abuzar Haroon

  3. Genome Research chair, Department of Biochemistry, College of Science, King Saud University, PO Box 2455, Riyadh, 11451, Saudi Arabia

    Mohamed Elrobh, Mohammed S. Alanazi & Mohammad D. Bazzi

  4. Department of Basic Sciences, College of Science and Health Professions, King Saud Bin Abdulaziz University for Health Sciences, PO Box 3660, Riyadh, 11426, Saudi Arabia

    Wajahatullah Khan

  5. Biochemistry Department, Faculty of Science, Ain-Shams University, Abbassia, 11381, Cairo, Egypt

    Mohamed Elrobh

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  1. Ajamaluddin Malik
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  2. Abuzar Haroon
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Correspondence to Ajamaluddin Malik.

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Malik, A., Haroon, A., Jagirdar, H. et al. Spectroscopic and thermodynamic properties of recombinant heat shock protein A6 from Camelus dromedarius . Eur Biophys J 44, 17–26 (2015). https://doi.org/10.1007/s00249-014-0997-2

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  • DOI: https://doi.org/10.1007/s00249-014-0997-2

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