Cloning and Expression of a Novel Gene Encoded Thermostable Archaeal Aldolase Class Ii from Natural Sample

Santi Nurbaiti¹, Akhmaloka¹*, Suharti^1,2, Nishia Waya Meray^1,3

¹Biochemistry Research Group, Faculty of Mathematics and Natural Sciences, Institute Teknologi Bandung, Jalan Ganesha 10, Bandung, 40132 Indonesia ²faculty of Pharmacy, Universitas 17 Agustus 1945 (UTA’45) Jakarta, Jl. SunterPermai Raya, Sunter Agung Podomoro, Jakarta, Indonesia ³Department of Oil and Gas Processing Technology, Sekolah Tinggi Teknologi Minyakdan Gas, Jl. Soekarno Hatta KM 8, KarangJoang, Balikpapan, Kalimantan Timur, Indonesia  

ABSTRACT: A novel class II Aldolase was isolatedthroughmetagenomic approach from DomasCrater, West Java, Indonesia. Sequenceanalysis of the enzymewas highly homolog to archaealribulose-5-phosphate 4-epimerase from unculturedAcidilobussp.with percent identityof 63%. Homological analysis of the protein shows that the protein sequence contains all conservedmotifs of aldolase class II. The enzyme shows Zn²⁺ binding, polypeptide binding, and active sites as other aldolase’s. Phylogenetic analysis of the enzyme showed that the enzyme makes a different branch closed to ribulose-5-phosphate 4-epimerase of unculcuredAcidilobus sp. The genewas expressed in E coli as a host, and produced26kDa of protein. Further analysis of the enzymeshowed that the enzyme is thermostable. In addition, the enzymewas purified through Ion Metal Affinity Chromatography and it showed as single band with the homogeneity at around 96%.

KEYWORDS: Keywords; aldolase; metagenom; thermostable; cloning

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