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Protein - Peptide Interaction Analysis
Application Note NT-MO-007
Binding of Histone peptides to Chromatin assembly factor I
(CAF-I) p48 subunit
Wei Zhang
University of Cambridge, Department of Biochemistry, Cambridge, UK
Abstract
p48, the small subunit of chromatin assembly
factor 1 (CAF-1), is a member of a highly
conserved subfamily of WD-repeat proteins.
There are at least two members of this
subfamily in human (p46 and p48). p48
copurifies with a chromatin assembly complex
(CAC), which contains the three subunits of
CAF-1 (p150, p60, p48) and the Histones H3
and H4, and promotes DNA replication-
dependent chromatin assembly.
In this study we analyze the binding of H3 and
H4 peptides to p48 using MicroScale
Thermophoresis (MST). The study also
highlights the high content information of the
MST measurements as one important benefit
of MicroScale Thermophoresis.
Introduction
Five major classes of histones exist: H1/H5, H2A,
H2B, H3, and H4. Histones H2A, H2B, H3 and H4
are known as the core histones, while histones H1
and H5 are known as the linker histones. Two of
each of the core histones assemble to form one
octameric nucleosome core particle, and 147 base
pairs of DNA wrap around this core particle 1.65
times in a left-handed super-helical turn. In
contrast the linker histone H1 binds the
nucleosome at the entry and exit sites of the DNA,
thus locking the DNA into place and allowing the
formation of higher order structure. Histone H5
performs the same function as histone H1, and
replaces H1 in certain cells. Histone proteins also
play essential structural and functional roles in the
transition between active and inactive chromatin
states.
Fig. 1 Crystal structure showing the octameric core
nucleosome particle consisting of two of each H2A, H2B, H3
and H4 Histones and a 147 base pairs DNA wrapped around
this core particle (PDB id: 1kx5)
Chromatin Assembly Factor-1 (CAF-1) assembles
newly synthesized histones H3/H4 into DNA in the
first step of nucleosome assembly. Accordingly in
human cells, CAF-1 is complexed to newly
synthesized and acetylated histones H3 and H4.
Human CAF-1 consists of three subunits: p150,
p60 and p48. The small CAF-1 subunit p48 is a
member of a highly conserved subfamily of WD-
repeat proteins. p48 proteins are involved as
histone escorts in a multistep process of
cytoplasmic histone acetylation, the assembly of
histones into chromatin, and the subsequent
deacetylation of incorporated histones. p48
copurifies with a chromatin assembly complex
(CAC), which contains the three subunits of CAF-1
(p150, p60, p48) and H3 and H4. There are at
least two human homologs which are called
