Erythrocyte core particles

The inverse of the above experiments gave similar results (Whitlock and Stein, 1978). Trypsin-digested histones removed from HeLa core particles can subsequently fold DNA, although DNase I digests the resulting particles more rapidly than the untreated ones. Parallel experiments were performed for chicken erythrocyte core particles (Lilley and Tatchell, 1977). In all cases it could be concluded that it is the trypsin-insensitive carboxy-terminal regions of the histones which are responsible for the folding of the DNA in the nucleosome. 

Fig. 1. Time course study of poly(ADP-ribosyl)ation of chicken erythrocyte core particles (O—O), calf thymus native chromatin (A—A), and HI-depleted chromatin (A—A) by purified calf thymus poly(ADP-ribose) polymerase at 30°C. Poly(ADP-ribose) polymerase (2 Mg/OD unit), which was purified free of its DNA according to Zahradka and Ebisuzaki [15], was incubated with various chromatin preparations at 200 nM NAD. At various times, the reaction was stopped by the addition of 10% TCA (Cl3AcOH)/2% PPi, and the activity was determined according to Aubin et al. [8, 9]. — represents the endogenous activity of the poly(ADP-ribose) polymerase found on calf thymus native chromatin...

Interestingly poly(ADP-ribosyl)ation of chicken erythrocyte core particles resulted in the dissociation of nucleosomal DNA from the histone octamer (Fig. 3c,f). The dissociation coincided with the generation of the hyper(ADP-ribosyl)ated forms of histone H2B (Fig. 2). This result is noteworthy because on exposing hepatoma cells to DMS, histone H2B becomes hyper(ADP-ribosyl)ated [18]. The dissociation of nucleosomal DNA from the histone octamer after poly(ADP-ribosyl)ation explains why nucleosomal DNA becomes more accessible to micrococcal nuclease [19]. The increased accessibility of nucleosomal core DNA caused by poly(ADP-ribosyl)ation could explain in part the increased accessibility of DNA repair patches to micrococcal nuclease observed during the early phase of DNA repair [20]. 

We have studied the interaction between poly(ADP-ribose) and purified chicken erythrocyte core particles which had been prepared according to Erard et al. [23] or Lutter [24]. When core particles and poly(ADP-ribose) were incubated and then separated by velocity sedimentation (Fig. 6), some interaction between polymer and core particles was observed. 

Fig. 6. Interaction between poly(ADP-ribose) and chicken erythrocyte core particles. [ P]-poly-(ADP-ribose) was synthesized by purified poly(ADP-ribose) polymerase incubated with calf thymus polynucleosomes and the polymer was extracted as described by Miwa et al. [26]. Core carticles and the polymer were allowed to interact and then centrifuged on a 5-20% (w/v) isokinetic sucrose gradient containing 40 vM NaCl, 10 xM Iris, 0.2 mM EDTA at pH 7.4. Centrifugation was for 14 h at 40,000 rpm (4°C) in a SW41Ti Beckman rotor...

Fig. 6. Purified recombinant HMGA proteins bind to four regions of DNA on random sequence nucleosome core particles. Panel A The results of EMSA gel assays in which increasing concentrations of either purified nonhistone HMGN2 (a.k.a., HMG-17, which binds to two sites on nucleosome core particles) or recombinant human HMGAla protein were bound nucleosome core particles isolated from chicken erythrocytes [57]. Panel B Two different views of the nucleosome taken from the X-ray structure of Luger et al. [119] showing the sites of binding of HMGA proteins (dashed circles) determined by DNA foot-printing analyses and other techniques (see text for details).

Simpson and Shindo (1980) have compared the P-NMR spectra at 109.3 MHz of three 145-bp double-stranded DNAs (Fig. 4), namely, poly(dGdC)-poly(dGdC), poly(dAdT) poly(dAdT), and random-sequence chicken erythrocyte DNA obtained from nucleosome core particles. In... 

Fig.l3. P-NMR spectra at 109.3 MHz ofchicken erythrocyte nucleosome core particles in 5 mA/Tris-HQ, pH 8, alone (upper spectrum) and in the presence of increasing concentrations ofMnG2(seeFig. 14). 

Different objects may be used as the sacrificial core to prepare hollow capsules, e.g. particles of polymers e.g., melamine-formaldehyde resin, poly(sodium styrenesulfonate) (PSSS) ], inorganic compounds [e.g., CaCOg or SiOa ), metal nanoparticles, or even bio-objects such as erythrocytes or platelets. Multilayers can be also deposited directly on the nano- or microcrystals of drugs. 

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