Abstract
Nucleic acid binding proteins have important roles in DNA and RNA packaging, stabilisation and repair, and in gene regulation, and they are therefore essential for all organisms. All of the known hyperthermophiles have at least one DNA sequence encoding for the Alba proteins. The Alba proteins are small (approximately 10 kDa), DNA-binding, basic proteins that appear to partly compensate for the lack of histones in the archaea Aeropyrum pernix and other hyperthermophiles. Two sequences of these potential histone counterparts, the Alba proteins, were identified in the Aeropyrum pernix genome (APE1832.1 and APE1823). By using a wide range of experimental techniques and by examining several combinations of expression systems the expression of recombinant Alba1 and Alba2 proteins was optimized. Co-expression of both of the Alba proteins was needed when isolating recombinant Alba2. The purification of both recombinant Alba1 and Alba2 His-tagged proteins were simplyfied in satisfactory yield. The electrophoretic mobility shift assay demonstrated the ability of the Alba1 and Alba2 proteins from Aeropyrum pernix to bind DNA. (doi: 10.5562/cca1772)
Highlights
Archaea are one of three presently recognised phenotypical domains[1] and they are widely known for their ability to inhabit extreme habitats
The Alba proteins were expressed as GSTtagged products by applying heterologous expression in E. coli, which resulted in relatively low protein yields due to low protein solubility
To increase the protein yield with the use of these His-tagged expression systems, we investigated the effects of different growth media, growth temperatures and isopropyl β-D-thiogalactoside (IPTG) concentrations
Summary
Archaea are one of three presently recognised phenotypical domains[1] and they are widely known for their ability to inhabit extreme habitats. Archaeal nucleic acids are usually stabilised with a combination of different stabilisation mechanisms, which include higher GC content (%) in their RNA molecules, binding of cations and polyamines, covalent modifications of mainly RNA molecules, compact tertiary structures of nucleic acids, binding of thermostable proteins, and efficient repair systems.[2] Besides histones, the most widespread DNAbinding proteins are the Alba proteins, which are present in all known hyperthermophiles.[3] Aeropyrum pernix is a hyperthermophilic crenarchaeon, and it is a strictly aerobic organism with an optimal growth temperature of around 90 °C. A. pernix contains no known histones.[4,5] The histone function is believed to be carried out by other proteins, such as the 10 kDa small basic proteins in archaea (the Alba proteins) and other basic protein families (e.g. 7 kDa small basic proteins).[6,7,8,9,10]
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