Methods in Molecular Biology • 16 Enzymes of Molecular Biology

112 Pingoud, Alves, and Geiger Table 2 Structural Properties of Selected Restriction Enzymes Subunit Enzyme Amino acid res. Mol ...

Restriction Enzymes 113 there is no sequence similarity over an extended stretch of amino acids. Lauster (52), however, has dete ...

114 Pingoud, Alves, and Geiger linear diffusion (~ I I .~- ",,. (~ (k') specific non-1~ / specific 1~ binding binding cleavage c ...

Restriction Enzymes 115 accelerates the rate of the cleavage reaction (64-67) by lowering the apparent K m. Linear diffusion alo ...

116 Pingoud, Alves, and Geiger © °1 tO e- Q; r, r,~ p- q._ ._,ooo --."~ ~ .-~ooo, 6~ .... ~' ~ ~-~= z~6 ~= ~a ~'~ ~<~ ~D er~ ...

Restriction Enzymes 117 oligodeoxynucleotide substrates have much higher K m values (78, 83, 84, 8 7, 92, 93), presumably becaus ...

118 Pingoud, Alves, and Geiger sequences adjacent to the recognition site have shown for EcoRI that AT base pairs enhance and GC ...

Restriction Enzymes 119 scissile phosphodiester bond, rather than with an involvement of a covalent enzyme intermediate (115). I ...

120 Pingoud, Alves, and Geiger the recognition site is used by the enzyme to differentiate between spe- cific and nonspecific DN ...

Restriction Enzymes 121 2.4.1. Methylation Methylation of critical residues of the recognition sequence in gen- eral prevents a ...

122 Pingoud, Alves, and Geiger conditions of elevated pH and low ionic strength, EcoRI cleaves DNA not only within the canonical ...

Restriction Enzymes 123 or d(TCTCCGGTT) by MspI (153) may be owing to duplex formation via the palindromic -CCGG- core sequence, ...

124 Pingoud, Alves, and Geiger to p-mercuribenzoate, N-ethyl maleimide, 5,5'-dithiobis (2-nitro- benzoic acid), and iodoacetate, ...

Restriction Enzymes 125 cysteine residues), 0.01-0.1% (w/v) Triton X-100, Tween, Lubrol, or other detergents, as well as 0.1 mg/ ...

126 Pingoud, Alves, and Geiger Table 4 Resolution of Different Gel Systems for Electrophoresis of DNA Fragments Separation range ...

Restriction Enzymes 127 quantities of DNA can be seen on inspection of the gel under UV light. Silver staining allows the detect ...

128 Pingoud, Alves, and Geiger gels, whereas complete EcoRV-~,-fragments (ranging from 270-5770 bp) require separation on at lea ...

Restriction Enzymes 129 3.4.2. Ionic Strength The accuracy and activity of restriction enzymes are strongly affected by the ioni ...

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o o ,,~ ,.-,o o o v'SUe~v= ~= Ot"4 o ~= [L e~ (^00) ~t3 c4 4- 4- + oo t"qo ~ o 0 r.~ ~"~ t '~ I I I I +< r..) I + I =,>" ...