Zinc Metalloproteins Involved in Replication and Transcription
David P. Giedroc, Kathleen M. Keating, Craig T.
Martin, Kenneth R. Williams, and Joseph E. Coleman
J. Inorg. Biochem.28, 155-169, 1986.
RNA polymerase (RPase) from E. coll contains two tightly
incorporated Zn(II) ions, while the monomeric RPase from bactenophage T7 does
not contain zinc and does not require Zn(II) in the assay. One of the two
Zn(II) ions can be differentially removed from E coli RPase with p-hydroxymer-curiphenylsulfonate
(PMPS) combined with EDTA and thiol. The resultant Zn1 or ZnA RPase shows no
alteration in transcription initiation and elongation rate from -specific
promoters. Biosynthesis of a Co2
RPase and formation of CoA RPase by similar
treatment shows the tetrahedral-type Co(II) d-d absorption bands to be associated only
with the Co(II) at the A site with maxima at 760 ( e= 800), 710 ( e= 900), 602 (
e= 1500), and 484 (e = 4000) nm. Sulfur to Co(II) charge transfer bands are
present at 350 ( e= 9600) and 370 ( e= 9500) nm. The absorption characteristics
strongly suggest that the A site is a tetrathiolate site. While DNA polymerases
do not in general appear to contain zinc, gene 32 protein (g32P) from
bactenophage T4, an accessory protein essential for DNA replication and
recombination and translational control in the T4 life cycle, is a Zn(II)
metalloprotein and contains 1 gram atom of tightly incorporated Zn(II) PMPS
displaces the zinc by reacting with three SH groups. Apo-g32P shows markedly
altered DNA binding properties. Co(II) substitution gives a protein with
intense d-d transitions typical of
a tetrahedral Co(II) complex with absorption maxima at 680 ( e= 480), 645 ( e=
660), 605 ( e= 430), 355 ( e= 2250), and 320 ( e= 3175) nm. The data support a 3
Cys, 1 His coordination site located in the middle of the DNA binding domain of
g32P. Data thus far suggest that the Zn(II) binding sites in multisubunit RNA
polymerases and in accessory proteins involved in polynucleotide biosynthesis
are more likely to play structural or allosteric (regulatory) roles rather than
directly participating in catalysis.
