Cloning, expression, and biochemical characterization of a new histone deacetylase-like protein from Thermus caldophilus GK24.
Histone deactylases (HDACs) are members of an ancient enzyme family found in eukaryotes as well as in prokaryotes such as archaebacteria and eubacteria. We here report a new histone deacetylase (Tca HDAC) that was cloned from the genomic library of Thermus caldophilus GK24 based on homology analysis with human histone deacetylase1 (HDAC1). The gene contains an open reading frame encoding 375 amino acids with a calculated molecular mass of 42,188 Da and the deduced amino acid sequence of Tca HDAC showed a 31% homology to human HDAC1. The Tca HDAC gene was over-expressed in Escherichia coli using a Glutathione-S transferase (GST) fusion vector (pGEX-4T-1) and the purified protein showed a deacetylase activity toward the fluorogenic substrate for HDAC. Moreover, the enzyme activity was inhibited by trichostatin A, a specific HDAC inhibitor, in a dose-dependent manner. Optimum temperature and pH of the enzyme was found to be approximately 70 degrees C and 7.0, respectively. In addition, zinc ion is required for catalytic activity of the enzyme. Together, these data demonstrate that Tca HDAC is a new histone deacetylase-like enzyme from T. caldophilus GK24 and will be a useful tool for deciphering the role of HDAC in the prokaryote and development of new biochemical reactions.
Song
Young Mi
YM
Chemical Genomics Laboratory, Department of Biotechnology, College of Engineering, Yonsei University, Seoul 120-749, Republic of Korea.
Kim
You Sun
YS
Kim
Dooil
D
Lee
Dae Sil
DS
Kwon
Ho Jeong
HJ
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D015870:Gene Expression; D015723:Gene Library; D006655:Histone Deacetylases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000032:analysis / Q000302:isolation & purification; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013824:Thermus / Q000201:enzymology*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D006655:Histone Deacetylases