Cloning and sequencing of the gene coding for S-adenosylhomocysteine hydrolase in the thermophilic archaeon Sulfolobus solfataricus.
The gene from the thermophilic archaeon Sulfolobus solfataricus (Ss), encoding the S-adenosylhomocysteine hydrolase (AdoHcyHD), has been cloned. Two degenerate oligodeoxyribonucleotide (oligo) probes, synthesized on the basis of amino acid (aa) sequence of cyanogen bromide-peptide fragments of the purified protein, were used to screen a genomic library of Ss cloned into the pGEM7Zf(+) vector. The AdoHcyHD gene (adohcyhd) comprises 1254 nucleotides (nt) and encodes a polypeptide of 417 aa with a deduced molecular mass of 46 kDa, in good agreement with the value directly measured for the purified enzyme. The identity of more than 32% of the deduced aa sequence was confirmed by Edman degradation of peptides. Putative regulatory elements which are in good agreement with the archaeal promoter consensus sequences were identified in the flanking regions. Comparison of the aa sequences of AdoHcyHD from different sources shows a remarkable degree of conservation. Surprisingly, several aa residues, thought important in substrate binding and catalysis, show non-conserved replacements in Ss AdoHcyHD.
Porcelli
M
M
Istituto di Biochimica delle Macromolecole, Facoltà di Medicina e Chirurgia, Seconda Università degli Studi di Napoli, Naples, Italy.
Cacciapuoti
G
G
Fusco
S
S
Bertoldo
C
C
De Rosa
M
M
Zappia
V
V
D043383:Adenosylhomocysteinase; D000595:Amino Acid Sequence; D001483:Base Sequence; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D005798:Genes, Bacterial; D006867:Hydrolases / Q000235:genetics*; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D016944:Sulfolobus / Q000201:enzymology* / Q000235:genetics
D004269:DNA, Bacterial; D006867:Hydrolases; D043383:Adenosylhomocysteinase