Isolation of amino-terminal fragment of lactose repressor necessary for DNA binding.
lac repressor can be dissected by trypsin into a homogenous tetrameric core (accounting for residues 60 to 347), carrying inducer binding activity, and the monomeric amino-terminal peptides ("headpieces") accounting for residues 1 to 59 and 1 to 51, respectively. This restriction of the action of trypsin on lac repressor is obtained in 1 M Tris-HCl (pH 7.5)-30% in glycerol at 25 degrees C since only the peptide bonds at lysine-59 and to a lesser extent after at arginine-51 are cleaved under these conditions. The headpieces can be purified by gel filtration. They have ordered secondary structure as revealed by circular dichroism studies. The monomeric headpieces show the relatively weak binding to nonoperator DNA but not the highly specific and strong binding to operator DNA typical for tetrameric lac repressor.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism; D001665:Binding Sites; D002942:Circular Dichroism; D003488:Cyanogen Bromide; D004247:DNA / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D007785:Lactose / Q000378:metabolism*; D009698:Nucleoproteins / Q000302:isolation & purification / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D011487:Protein Conformation; D014357:Trypsin
D000596:Amino Acids; D001426:Bacterial Proteins; D009698:Nucleoproteins; D010446:Peptide Fragments; D004247:DNA; D014357:Trypsin; D007785:Lactose; D003488:Cyanogen Bromide