Core complexes

The core unit of the chromatin, the nucleosome, consists of histones arranged as an octamer consisting of a (H3/ H4)2-tetramer complexed with two histone H2A/H2B dimers. Accessibility to DNA-binding proteins (for replication, repair, or transcription) is achieved by posttranslational modifications of the amino-termini of the histones, the histone tails phosphorylation, acetylation, methylation, ubiquitination, and sumoyla-tion. Especially acetylation of histone tails has been linked to transcriptional activation, leading to weakened interaction of the core complexes with DNA and subsequently to decondensation of chromatin. In contrast, deacetylation leads to transcriptional repression. As mentioned above, transcriptional coactivators either possess HAT activity or recruit HATs. HDACs in turn act as corepressors. 

Figure 37-2. RNA polymerase (RNAP) catalyzes the polymerization of ribonucleotides into an RNA sequence that is complementary to the template strand of the gene. The RNA transcript has the same polarity (5 to 3 ) as the coding strand but contains L) rather than T. E coli RNAP consists of a core complex of two a subunits and two p subunits (P and p ). The holoen-zyme contains the 0 subunit bound to the ajPP core assembly. The co subunit is not shown. The transcription "bubble" is an approximately 20-bp area of melted DNA, and the entire complex covers 30-75 bp, depending on the conformation of RNAP.

It is also interesting to note that only a fraction of PS II membrane protein forms a stable monolayer structure and the rest of them fall into the water subphase. This can be seen directly by the naked eye during the compression. Furthermore, if we use the total amount of PS II membrane protein to calculate the average particle size from the n-A curve, we obtain an area of about 200 nm. This value is very small when compared with that of the PS II core complex (320 nm, as discussed in the subsequent section), which is a smaller subunit of the PS II membranes. A PS II membrane fragment contains PS II core complex and several LHC II proteins, and is much larger in size than a PS II core complex... 

In the case of PS II membrane proteins, as discussed above, the hydrophobic and hydrophilic pairs of attached lipids can partially support the protein complex at the air-water interface, despite their large size and density. However, in the case of PS II core complex, the detergent strips the attached lipids and some extrinsic proteins. The remaining protein complex is water soluble. It is very difficult to prepare a stable monolayer of water-soluble proteins with the Langmuir method. Indeed, it is hard to directly prepare a stable monolayer of PS II core complex because of its water solubility as well as density. One possible solution is to change the density and ionic strength of the subphase [9]. 

We studied the surface pressure area isotherms of PS II core complex at different concentrations of NaCl in the subphase (Fig. 2). Addition of NaCl solution greatly enhanced the stability of monolayer of PS II core complex particles at the air-water interface. The n-A curves at subphases of 100 mM and 200 mM NaCl clearly demonstrated that PS II core complexes can be compressed to a relatively high surface pressure (40mN/m), before the monolayer collapses under our experimental conditions. Moreover, the average particle size calculated from tt-A curves using the total amount of protein complex is about 320 nm. This observation agrees well with the particle size directly observed using atomic force microscopy [8], and indicates that nearly all the protein complexes stay at the water surface and form a well-structured monolayer. 

FIG. 2 The surface pressure-area isotherms of PS II core complex with different concentrations of salt in the subphase. Subphase, lOmM tris-HCl, pH 8.0, 2mM sodium ascorbate and concentrations of 100, 200, and 500mM NaCl. Temperature, 23.0 0.5°C. 

Our studies on the surface pressure-area isotherms of MGDG and the mixture of PS II core complex and MGDG indicate the presence of both PS II core complex and MGDG in the monolayer. MGDG molecules diluted the PS II core complex concentration in the monolayer. MGDG lipid functions as a support for the protein complex and the resulting mixture forms higher-quality films than PS II core complex alone. 

The A F-A isotherm of PS II core complex is rather different from that of PS II membrane (Fig. 4). The surface potential of a monolayer of PS II core complex increases slightly as the molecular area is compressed from 600 to about 150nm, while surface pressure changes from 5 to 35mN/m. Further compression results in a sharper increase in surface potential. The surface potential starts to decrease only after the surface area is compressed to about 80 nm or surface pressure becomes larger than 40mN/m. This is consistent with the previous discussion that PS II core complexes form a more ordered monolayer structure at relatively high surface potential and will not form multilayered... 

FIG. 4 The surface pressure-area (tt-A) and surface potential-area (A V-A) isotherms of PS II core complex particles. 

Phan, L., Zhang, X., Asano, K., Anderson, J., Vomlocher, H. P., Greenberg, J. R., Qin, J., and Hinnebusch, A. G. (1998). Identification of a translation initiation factor 3 (eIF3) core complex, conserved in yeast and mammals, that interacts with eIF5. Mol. Cell. Biol. 18, 4935-4946. 

APS represent an attractive novel way of stabilizing membrane proteins under conditions where they can be exposed to detergent-free media. In the present work, we report some preliminary experiments aimed at complexing with APS a photosynthetic PS2 reaction center core complex, that from the thermophilic cyanobacterium Thermosynechococcus elongatus. 

Figure 3. Size exclusion chromatography analysis of AP-trapped vs. detergent-solubilized FS2 core complexes.

Figure 4. Oxygen evolving activity of AP-trapped v.v. detergent-solubilized PS2 core complexes.

PS1 and PS2 core complexes were solubilized by 1% SDS and separated by SDS-PAGE using a 12 % gel containing 6 M urea according to[Schagger and von Jagow 1987],... 

Meyer, B., et. al., Hepatitis B virus X-associate protein 2 is a subunit of the unliganded aryl hydrocarbon receptor core complex and exhibits transcriptional enhancer activity, Mol. Cell. Biol., 18, 978, 1998. 

Nakazato K, Ichimwa T, Mayanagi K, Ishikawa T, Inoue Y. Atomic force microscopy of two-dimensional crystal of photosystem II core complex. Plant Cell Physiol 1998 (Suppl) 39 S12. 

In experiments performed with chromatin or core particles depleted of histones except for H3 and H4 (Stockley and Thomas, 1979), two complexes were obtained, one containing an octamer and one a tetramer of H3 and H4 per 140 base pairs of DNA. The physical properties of the two core complexes were similar to those observed by... 

Masison, C. a., Radonovich, M., Phan, L., Clayton, J., He, H., Beady, J. N., Hinnebusch, a. G., Asano, K. Saccharomyces cerevisiae protein Pci8p and human protein eIF3e/Int-6 interact with the eIF3 core complex by binding to cognate eIF3b subunits. J. Biol. Chem. 2001, 276, 34948-34957. 

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