Complete nucleotide sequence of pSTK1, a cryptic plasmid from Bacillus stearothermophilus TK015

Abstract

The complete nucleotide sequence of pSTK1, a cryptic plasmid isolated from B. stearothermophilus TK015, has been determined, pSTK1 has been shown to be 1883 bp in length and contain three open reading frames (ORFs), one of which has a helixturn-helix motif typical of DNA-binding proteins. Also identified was a region that can form an extensive secondary structure, which would show a high degree of similarity to palA, an origin for minus strand elongation in roiling circle replication. INTRODUCTIONThe 1.9-kbp cryptic plasmid pSTK1 was originally isolated from B. stearothermophilus TK015, and it was stably maintained in its host up to 70 °C without changing its copy number. The thermostable nature of pSTK1 was inherited by plasmid pSTE33, which is a shuttle vector constructed from pSTK1, plasmid pUC19, and a thermostable kanamycin-resistance marker (Narumi et al., 1993). The thermostability and host range of pSTE33 make it useful for cloning experiments using B. stearothermophilus as a host.The part of pSTE33 nucleotide sequence corresponding to pUC19 (YanischPerron et al., 1985) and the thermostable kanamycin nucleotidyltransferase gene (Liao et al., 1986) is known. This paper describes 1) the complete nucleotide sequence of pSTK1, which is the key to the thermostable replication of pSTE33 in B. stearothermophilus, and 2) homologies of pSTK1 with other plasmids.

Genetic engineering of Geobacillus spp.

“…plasmids have been sequenced and described. These plasmids are pSTK1, pGS18, pGTG5, and pGTD7 (Kananavičiūtė et al, 2014;Nakayama et al, 1993;Stuknyte et al, 2008). Plasmids pBST1 (Taylor et al, 2008) and pTB19 (Oskam et al, 1991;Van der Lelie et al, 1989) have only been partially sequenced, while 10 more Geobacillus sp.…”

Regulation, replication, and integration functions of the Vibrio cholerae CTXφ are encoded by region RS2

“…rstA2 is predicted to encode a polypeptide of 359 amino acids. Database searches revealed that this protein is similar to a hypothetical polypeptide (ORF3) in pSTK1, which is a cryptic plasmid of Bacillus stearothermophilus (Nakayama et al, 1993). RstA contains a Walker 'type A' motif (A /G XXXXGKT; Walker et al, 1982) located 52 amino acid residues from the C-terminus, suggesting that this protein may hydrolyse an nucleoside triphosphate.…”

Nucleotide sequence and characterization of the cryptic Bacillus thuringiensis plasmid pGI3 reveal a new family of rolling circle replicons

“…The resulting construct (pGI301s) was unable to replicate in B. thuringiensis and B. subtilis, suggesting that ORF6 corresponded to the bona fide replication protein of pGI3. As mentioned above, ORF6 showed 57.4% similarity to one of the three ORFs (ORF3) present in pSTK1, a B. stearothermophilus plasmid (36). The alignment of the two proteins is shown in Fig.…”

“…Extensive database searches with the predicted products of these ORFs did not reveal any similar proteins, with the notable exception of ORF6 (see below), which showed 57.4% similarity to an ORF (ORF3) of pSTK1, a cryptic B. stearothermophilus plasmid (36). Database searches were also performed with the pGI3 DNA sequence, spliced into 500-bp fragments (with 250-bp overlap), using the GCG-BLAST program.…”