Cloning and characterization of the Hpall methylase gene

Abstract

The Hpall restriction-modification system from Haemophilus parainfluenzae recognizes the DNA sequence CCGG. The gene for the Hpall methylase has been cloned into E. coli and its nucleotide sequence has been determined. The DNA of the clones is fully protected against cleavage by the Hpall restriction enzyme in vitro, indicating that the methylase gene is active in E. coli. The clones were isolated in an McrA- strain of E. coli; attempts to isolate them in an McrA+ strain were unsuccessful. The clones do not express detectable Hpall restriction endonuclease activity, suggesting that either the endonuclease gene is not expressed well in E. coli, or that it is not present in its entirety in any of the clones that we have isolated. The derived amino acid sequence of the Hpall methylase shows overall similarity to other cytosine methylases. It bears a particularly close resemblance to the sequences of the Hhal, BsuFl and Mspl methylases. When compared with three other methylases that recognize CCGG, the variable region of the Hpall methylase, which is believed to be reponsible for sequence specific recognition, shows some similarity to the corresponding regions of the BsuFl and Mspl methylases, but is rather dissimilar to that of the SPR methylase. © 1990 Oxford University Press.