Question

What aspect of heparin allows it to bind effectively to the RNA polymerase active site?

Answer 1

Heparin is a very potent and specific inhibitor of
RNA polymerases in that it interferes with the initia-
tion of transcription. Therefore the term affinity
chromatography is justified to describe the binding of
the enzyme to the immobilised inhibitor. Affinity
chromatography of E. coli RNA polymerase on
matrix-bound heparin provides the holo enzyme in
high yield. This preparation is twice as active as other
enzyme preparations. The higher specific activity
may have several reasons. First of all we believe that
the unusually high content of factor cr of the enzyme
purified on heparin-Sepharose is responsible for the
increased specific activity. RNA polymerase which
is prepared by a common procedure shows 30- 50 %
deficiency of factor cr in comparison to the other sub

units. By addition of factor (T to such an enzyme

the translation of native DNA can be stimulated

very effectively. Furthermore probably enzymically

inactive RNA polymerase does not bind tightly to

heparin-Sepharose as was shown with rifampicin-

Sepharose by other authors [16]. Lastly heparin also

inhibits ribonucleases. These may also have a pro-

nounced but different affinity for the heparin-sepha-

rose and can thus be removed quantitatively. Degrada-

tion of the reaction products of RNA polymerase

is therefore very unlikely.