ABSTRACT
When searching for the site-specific endonucleases in several strains of Phormidium we made the following observations. Among the 16 strains that originated from 15 species of Phormidium, 12 produced one or more restriction enzymes, of which two produced the highly thermophilic restriction endonucleases PtaI and PpaAII with their optimum activity at 65-80 degrees C, which is far above the lethal temperature for the host microorganism (40 degrees C). These two temperature-resistant enzymes are isoschizomers of known BspMII and TaqI endonucleases, respectively. The presence of the thermophilic TaqI isoschizomer does not seem to play any role in the mesophilic host microorganism, which does not even contain an active cognate methyltransferase. Among the remaining 10 strains, six produced isoschizomers of endonucleases which we first described in cyanobacteria, namely: PfaAII (NdeI), PinBII and PtaI (BspMII), PlaAII (RsalI), PpaAII, PpeI (ApaI). Two enzymes, PauAII (AhaIII) and PfaAII (NdeI), belong to a group of a very rarely occurring isoschizomers. Out of 21 cyanobacterial endonucleases investigated by us, four were active in a wide range of temperatures (from 15 to 60 degrees C) which also extended the optimal growth temperature of the hosts. We assume that our observation on the presence of temperature-resistant restriction enzymes in mesophilic hosts supports the idea of horizontal gene transfer. Restriction modification systems may be an excellent tool for investigation of that phenomenon.
Subject(s)
Cyanobacteria/enzymology , DNA Restriction Enzymes/chemistry , DNA Restriction Enzymes/metabolism , Cyanobacteria/classification , Cyanobacteria/growth & development , Deoxyribonucleases, Type II Site-Specific/metabolism , Enzyme Stability , Temperature , Thermodynamics , Thermus/enzymologyABSTRACT
A novel type II restriction endonuclease UnbI was isolated from an unidentified psychrofilic bacterial strain from Antarctica. UnbI recognizes and cleaves the sequence 5'-GGNCC-3', producing 5 nucleotide long sticky ends. In this respect it differs from its neoschizomer Sau96I and all other restriction enzymes recognizing this sequence. UnbI has a relatively low temperature optimum of 15 degrees C to 20 degrees C and its activity is completely inhibited by inorganic phosphate.
Subject(s)
Deoxyribonucleases, Type II Site-Specific/antagonists & inhibitors , Deoxyribonucleases, Type II Site-Specific/isolation & purification , Gram-Negative Aerobic Rods and Cocci/enzymology , Antarctic Regions , Base Sequence , Binding Sites , Cold Temperature , DNA/genetics , DNA/metabolism , Deoxyribonucleases, Type II Site-Specific/metabolism , Gram-Negative Aerobic Rods and Cocci/isolation & purification , Molecular Sequence Data , Phosphates/pharmacology , Substrate SpecificityABSTRACT
PpeI is a type II restriction endonuclease isolated from cyanobacterial strain Phormidium persicinum. The endonuclease PpeI, an isoschizomer of ApaI, recognizes the hexanucleotide sequence (5'-GGGCC/C-3') and cleaves, after the second C, producing four nucleotide 3'-cohesive ends.
Subject(s)
Cyanobacteria/enzymology , Deoxyribonucleases, Type II Site-Specific/isolation & purification , Bacteriophage lambda/genetics , DNA, Viral/metabolism , Deoxyribonucleases, Type II Site-Specific/metabolism , Hydrogen-Ion Concentration , Osmolar Concentration , TemperatureABSTRACT
We have isolated two restriction endonucleases, PinBI and PinBII, from the cyanobacterial strain Phormidium inundatum, and identified them as isoschizomers of AvaIII and BspMII, respectively.
