MED1 Glycosylase, a Tool for Detection of transition SNPs at CpG sites (CT-SNPs)

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Developed by:

  • Alfonso Bellacosa, MD, PhD

  • Introduction:

    Dr. Alfonso Bellacosa and colleagues of the Fox Chase Cancer Center have discovered MED1 (also known as MBD4), a novel human gene involved in the repair of mismatched bases.

    MED1 protein is a thymine and uracil glycosylase specific for G:T and G:U mismatches. Changes at CpG sites are the most frequent source of human genetic variation, as demonstrated by a recent survey by the group of Dr. Chakravarti on single nucleotide polymorphisms (SNPs) at candidate genes regulating blood pressure (Halushka et al. 1999).

    The availability of recombinant MED1 protein affords an easy three-step method to detect CT-SNPs, involving:

    • The formation of an heteroduplex containing a G:T mismatch.
    • Cleavage of the heteroduplex on the T-containing strand by the combined action of MED1 followed by incubation in hot alkali.
    • Separation of the cleaved molecules from the uncleaved molecules by various techniques, such as electrophoresis.
    • This three-step method is schematically represented in the attached Figure.

    This system requires a single CT-SNP probe for every CT-SNP to analyze. Multiple CT-SNP probes could be used to analyze a single PCR fragment containing multiple CT-SNPs, as long as the cleaved products generated by incubation with MED1 are of different length.

    A comparison of MED1 with other SNP detection tools is provided in the attached Table.

    Opportunity:

    Licenses are available for both commercial and research use.

    Publications:

    • Bellacosa A. et al. (1999) MED1, a Novel Human Methyl-CpG Binding Endonuclease, Interacts with the DNA Mismatch Repair Protein MLH1. Proc. Natl. Acad. Sci. USA 96:3969-3974
    • Petronzelli F. et al. (2000) Biphasic Kinetics of the Human DNA Repair Protein MED1 (MBD4), a Mismatch-Specific DNA N–Glycosylase. J. Biol. Chem. 275: 32422-32429
    • Petronzelli, F. et al. (2000) Investigation of the Substrate Spectrum of the Human Mismatch-Specific DNA Glycosylase MED1 (MBD4): the Fundamental Role of the Catalytic Domain. J. Cell. Physiol. 185: 473-480

    Patent Status:

    Applications are pending in the US, Europe, Canada and Australia.