trophoresis of HMG proteins routinely yields
20-40 bands, and 2-D electrophoresis of Hela
cell preparations results in more than 450 HMG
fractions, although in this case proteolytic artefacts cannot be excluded. The nuclear protein
spectrum of invertebrates, such as the Mediterranean fruit fly Ceratitis capitata or the nematode
Caenorhabditis elegans, have a comparably high
degree of complexity [57, 450]. In the absence of
information on the amino acid sequences, direct
comparisons of the non~histone proteins of vertebrates and invertebrates are impossible; however it is, remarkable that antibodies against
HMG-14 and HMG-17 of calf thymus show crossreactivity with the chromosomes of Drosophila.
In calf thymus, the largest fraction of the HMG
proteins is made up of the proteins numbered 1,
2, 14 and 17, the sequences of which are mostly
already known. Each nucleosome carries two
binding sites for HMG-14 or HMG-17. The
sequences of these two HMGs agree, for example, in the chicken at only 44 % of positions.
Compared with HMG-17, HMG-14 shows a much
higher rate of evolution: the human HMG-14 (98
amino acids) and that of the chicken (104 amino
acids) show only 51 % agreement, compared with
98 % agreement in the HMG-17 proteins [112].
HMG-l and HMG-2 from mammalian thymus
are identical in 79 % of their over 200 amino acids
[395]. The larger HMG-l and HMG-2 of calf thymus correspond to HMG-Tl and HMG-TI of the
rainbow trout Salrno gairdneri, and the smaller
HMG-14 and HMG-17 correspond to the HMGD and HMG-C; in addition, there are further
HMGs in fish that are not readily comparable
[51]. HMG-Tl and HMG-TI of the trout are each
encoded by four genes, whereas the human
HMG-14 and HMG-17 proteins are represented
by two multi-gene families, which in the case of
HMG-17 has 40-50 members [250]. The nucleoli
of all eukaryotes contain nucleolin, a multifunctional, 100-kDa protein that plays a key role
in rRNA maturation and the biogenesis of ribosomes. The sequence of 713 amino acids, determined via cDNA, shows no similarities to other
known proteins [251].
2.6 DNA Replication and Repair
DNA replication and repair in the eukaryotes has
been nowhere near as well investigated as in the
prokaryotes; the sparse comparative biochemical
2.6 DNA Replication and Repair
35
data that are available are restricted mainly to the
DNA polymerases, which in fact playa central
role in these processes. These enzymes link the
free 3'-OH group of an RNA or DNA primer to
the 5' -phosphate of a deoxynucleotide triphosphate. In the reactions of the polymerases, the
enzyme complex is either translocated to the next
nucleotide after each polymerization step (processive) or released (distributive). During the
replication of the chromosomal double helix, one
strand (the leading strand) is synthesized continuously, whereas the other (the lagging strand) is
synthesized only in sections (Okazaki fragments).
Thus, two different polymerase activities are
required for DNA replication: a highly processive
one for producing the leading strand, and one less
tightly bound polymerase that is released after
the replication of each Okazaki fragment. Five
types of DNA polymerase (a-E) are found in the
cells of mammals and other eumetazoans. Polymerases a and 0 take part in the replication of
chromosomal DNA, with the o-polymerase probably synthesizing the leading strand and the apolymerase the lagging strand [20,407, 427].
With only 10- 9 to 10- 12 errors per nucleotide per
generation, the reliability of DNA replication is
very high; in addition to the specificity of basepairing, this accuracy can be attributed to "proofreading", i.e. the excision of wrong nucleotides
by a polymerase-associated 3'-5'-exonuclease
activity [245]. The y-polymerase is responsible for
the replication of mitochondrial DNA, and the ~polymerase for the repair of chromosomal DNA
[478].
The a-polymerases are characterized by their
high molecular weight and by sensitivity to the
inhibitor N-ethylmaleimide (NEM); their function is the replication of chromosomal DNA. The
RNA primer required for the polymerase reaction
is synthesized by a primase that is always found
tightly associated with eukaryote a-polymerases.
The best-known animal a-polymerase primase is
that of Drosophila rnelanogaster. This is made up
of four subunits, a--O, of 182, 73, 60 and 50 kDa
respectively; polymerase activity is associated
with the 182-kDa subunit, primase activity with
the 60- and 50-kDa subunits; and no catalytic
activity is known for the 73-kDa subunit [92, 93].
The a-polymerases (replicases) of mammals
have molecular masses of up to 1300 kDa and a
highly complex structure that is not yet known in
detail [160, 224]. The a-polymerase of mammals
has no exonuclease activity but, in contrast, the
enzyme of Drosophila rnelanogaster exhibits
"cryptic" 3'-5' -exonuclease activity. The 0-
20-40 bands, and 2-D electrophoresis of Hela
cell preparations results in more than 450 HMG
fractions, although in this case proteolytic artefacts cannot be excluded. The nuclear protein
spectrum of invertebrates, such as the Mediterranean fruit fly Ceratitis capitata or the nematode
Caenorhabditis elegans, have a comparably high
degree of complexity [57, 450]. In the absence of
information on the amino acid sequences, direct
comparisons of the non~histone proteins of vertebrates and invertebrates are impossible; however it is, remarkable that antibodies against
HMG-14 and HMG-17 of calf thymus show crossreactivity with the chromosomes of Drosophila.
In calf thymus, the largest fraction of the HMG
proteins is made up of the proteins numbered 1,
2, 14 and 17, the sequences of which are mostly
already known. Each nucleosome carries two
binding sites for HMG-14 or HMG-17. The
sequences of these two HMGs agree, for example, in the chicken at only 44 % of positions.
Compared with HMG-17, HMG-14 shows a much
higher rate of evolution: the human HMG-14 (98
amino acids) and that of the chicken (104 amino
acids) show only 51 % agreement, compared with
98 % agreement in the HMG-17 proteins [112].
HMG-l and HMG-2 from mammalian thymus
are identical in 79 % of their over 200 amino acids
[395]. The larger HMG-l and HMG-2 of calf thymus correspond to HMG-Tl and HMG-TI of the
rainbow trout Salrno gairdneri, and the smaller
HMG-14 and HMG-17 correspond to the HMGD and HMG-C; in addition, there are further
HMGs in fish that are not readily comparable
[51]. HMG-Tl and HMG-TI of the trout are each
encoded by four genes, whereas the human
HMG-14 and HMG-17 proteins are represented
by two multi-gene families, which in the case of
HMG-17 has 40-50 members [250]. The nucleoli
of all eukaryotes contain nucleolin, a multifunctional, 100-kDa protein that plays a key role
in rRNA maturation and the biogenesis of ribosomes. The sequence of 713 amino acids, determined via cDNA, shows no similarities to other
known proteins [251].
2.6 DNA Replication and Repair
DNA replication and repair in the eukaryotes has
been nowhere near as well investigated as in the
prokaryotes; the sparse comparative biochemical
2.6 DNA Replication and Repair
35
data that are available are restricted mainly to the
DNA polymerases, which in fact playa central
role in these processes. These enzymes link the
free 3'-OH group of an RNA or DNA primer to
the 5' -phosphate of a deoxynucleotide triphosphate. In the reactions of the polymerases, the
enzyme complex is either translocated to the next
nucleotide after each polymerization step (processive) or released (distributive). During the
replication of the chromosomal double helix, one
strand (the leading strand) is synthesized continuously, whereas the other (the lagging strand) is
synthesized only in sections (Okazaki fragments).
Thus, two different polymerase activities are
required for DNA replication: a highly processive
one for producing the leading strand, and one less
tightly bound polymerase that is released after
the replication of each Okazaki fragment. Five
types of DNA polymerase (a-E) are found in the
cells of mammals and other eumetazoans. Polymerases a and 0 take part in the replication of
chromosomal DNA, with the o-polymerase probably synthesizing the leading strand and the apolymerase the lagging strand [20,407, 427].
With only 10- 9 to 10- 12 errors per nucleotide per
generation, the reliability of DNA replication is
very high; in addition to the specificity of basepairing, this accuracy can be attributed to "proofreading", i.e. the excision of wrong nucleotides
by a polymerase-associated 3'-5'-exonuclease
activity [245]. The y-polymerase is responsible for
the replication of mitochondrial DNA, and the ~polymerase for the repair of chromosomal DNA
[478].
The a-polymerases are characterized by their
high molecular weight and by sensitivity to the
inhibitor N-ethylmaleimide (NEM); their function is the replication of chromosomal DNA. The
RNA primer required for the polymerase reaction
is synthesized by a primase that is always found
tightly associated with eukaryote a-polymerases.
The best-known animal a-polymerase primase is
that of Drosophila rnelanogaster. This is made up
of four subunits, a--O, of 182, 73, 60 and 50 kDa
respectively; polymerase activity is associated
with the 182-kDa subunit, primase activity with
the 60- and 50-kDa subunits; and no catalytic
activity is known for the 73-kDa subunit [92, 93].
The a-polymerases (replicases) of mammals
have molecular masses of up to 1300 kDa and a
highly complex structure that is not yet known in
detail [160, 224]. The a-polymerase of mammals
has no exonuclease activity but, in contrast, the
enzyme of Drosophila rnelanogaster exhibits
"cryptic" 3'-5' -exonuclease activity. The 0-