Chemistry of Life

Molecular genetics, biochemistry

restriction enzmes

Bacterial restriction enzymes cut within the DNA molecule, so they are often called restriction endonucleases. A consensus sequence may be a short sequence of nucleotides that is found several times in the genome and is thought to play the same role in its different locations. Restriction enzymes usually have palindromic consensus sequences, which correspond to the site where they cut the DNA.

For example, the bacterium Hemophilus aegypticus produces a restriction endonucleases termed HaeIII that cuts DNA between the adjacent G and C wherever it encounters the sequence

5'GGCC3'
3'CCGG5'

HaeIII and AluI cut straight across the double helix producing "blunt" ends. However, many restriction enzymes cut in an offset fashion, generating an overlapping segment of single-stranded DNA. Such extensions are called "sticky ends" because they are able to form base pairs with any DNA molecule that contains the complementary sticky end. Such a union can be made permanent by another enzyme, DNA ligase, which forms covalent bonds along the backbone of each strand, producing a molecule of recombinant DNA (rDNA).

AluI and HaeIII produce blunt ends, while BamHI, HindIII, and EcoRI produce offset, “sticky” ends:

AluI................................................ 5’ … AGcutCT … 3’
Arthrobacter luteus............................. 3’ … TCcutGA … 5’

HaeIII........................... ................. 5’ … GGcutCC … 3’
Haemophilus influenzae....................... 3’ … CCcutGG … 5’

BamHI............................................. 5’ … GcutGATCC … 3’
Bacillus amyloliquefaciens................... 3’ … CCTAGcutG … 5’

HindIII............................................ 5’ … AcutAGCTT … 3’
Haemophilus influenzae .....................3’ … TTCGAcutA … 5’

EcoRI............................................... 5’ … GcutAATTC … 3’
Escherichia coli................................... 3’ … CTTAAcutG … 5’

The ability to produce recombinant DNA molecules revolutionized the study of genetics, and has proved important to biotechnology. Recombinant DNA technology has been utilized to produce human insulin (for diabetics), human factor VIII (for hemophilia A), and other useful proteins.

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. . . transcription begun 10/06/06