Cloning and characterization of the gene encoding PspPI methyltransferase from the Antarctic psychrotroph Psychrobacter sp. strain TA137. Predicted interactions with DNA and organization of the variable region.
The gene (pspPIM) encoding the PspPI DNA methyltransferase (MTase) associated with the PspPI restriction-modification (R-M) system (5'-GGNCC-3') of Psychrobacter species TA137 has been cloned and expressed in E. coli, and its nucleotide (nt) sequence has been determined. The coding region was 1248 nt in length and capable of specifying a 46826-Da protein of 416 amino acids (aa). The predicted sequence of the MTase protein displays ten sequence motifs characteristic of all prokaryotic m5C-MTases and shows the highest similarity to other MTases that methylate the GGNCC sequence, namely M . Eco47II and M . Sau96I. All three MTases methylate the internal cytosine within their recognition sequence. Sequence similarities between M . PspPI and its isospecific M . Eco47II and M . Sau96I as well as with four other m5C-MTases that methylate the related GGWCC sequence, namely M . SinI, M . HgiCII, M . HgiBI, M . HgiEI have been also found within the variable region of these proteins. On the basis of structural information from M . HhaI and M . HaeIII, several M . PspPI residues that are expected to interact with DNA can be predicted. Furthermore, an organization of the variable region of m5C-MTases into two segments exhibiting a pattern of conserved residues and a considerable degree of structural homologies is described.
Rina
M
M
Institute of Molecular Biology and Biotechnology, Enzyme Technology Division, Crete, Greece.
Caufrier
F
F
Markaki
M
M
Mavromatis
K
K
Kokkinidis
M
M
Bouriotis
V
V
D000595:Amino Acid Sequence; D000864:Antarctic Regions; D001426:Bacterial Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence / Q000235:genetics; D015257:DNA-Cytosine Methylases / Q000235:genetics* / Q000378:metabolism; D005798:Genes, Bacterial / Q000235:genetics*; D014644:Genetic Variation / Q000235:genetics*; D006088:Gram-Negative Aerobic Bacteria / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D015183:Restriction Mapping; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001426:Bacterial Proteins; D015257:DNA-Cytosine Methylases; C108719:PspPI DNA methyltransferase