Conformation of plasmid DNA and of DNA—histone chromatin-like complexes by laser light scattering

1979 ◽  
Vol 293 (1402) ◽  
pp. 303-313 ◽  
Keyword(s):  
Light Scattering ◽  
Molecular Mass ◽  
Plasmid Dna ◽  
Laser Light ◽  
Dynamic Properties ◽  
Mass Range ◽  
Laser Light Scattering ◽  
Biological Macromolecules ◽  
Experimental Conditions ◽  
Mass Size

Classical fluctuation and scattering theory has enabled the study of equilibrium properties such as molecular mass, size and intermolecular interactions of synthetic and biological macromolecules. Photon correlation and light beating spectroscopy, using laser radiation, allows the measurement of dynamic properties such as diffusion coefficients and, perhaps, the characterization of internal motion. The recently discovered plasmid DNA molecules are excellent objects of study, being monodisperse, in a convenient molecular mass range, and obtainable in supercoiled, relaxed circular and open linear conformations. Structural transitions can be studied in a variety of solvents and folding properties of DNA investigated. Chromatin can be reconstituted by interaction of DNA under well chosen experimental conditions with a selected group of histones, basic proteins of the nucleoprotein complex in the chromosome. The plasmid DNA-histone interaction leads to well-defined monodisperse chromatin-like complexes; their formation, composition and solution properties have been studied by laser light scattering and related physico-chemical methods.

scholarly journals Isolation and characterization of proteoglycans from human follicular fluid

1999 ◽  
Vol 340 (3) ◽  
pp. 613-620 ◽  
Author(s):  
Gitte V. ERIKSEN ◽  
Ingemar CARLSTEDT ◽  
Matthias MÖRGELIN ◽  
Niels ULDBJERG ◽  
Anders MALMSTRÖM
Keyword(s):  
Light Scattering ◽  
Molecular Mass ◽  
Heavy Chain ◽  
Follicular Fluid ◽  
Laser Light ◽  
Laser Light Scattering ◽  
Globular Domain ◽  
Human Follicular Fluid ◽  
Protein Core ◽  
Sds Page

Two proteoglycans differing in size and composition were isolated from human follicular fluid. The larger one of high density had a molecular mass of 3.0×106 Da, as determined by laser light-scattering, and was substituted with 15-20 chondroitin sulphate (CS) chains (Mr 60000-65000). Half of the CS disaccharides were 6-sulphated, whereas the remaining ones were non-sulphated. Digestion of the CS proteoglycan with chondroitinase ABC lyase, followed by SDS/PAGE, yielded a protein core of 600 to 700 kDa including substituted oligosaccharides, and a band of 70 kDa that was identified as the heavy-chain component of the inter-α-trypsin inhibitor (ITI). Western blotting of the CS proteoglycan showed that this had reactivity with antibodies raised against human versican. Electron microscopy (EM) of the CS proteoglycan also revealed a versican-like structure, with one globular domain at each end of a long extended segment substituted with CS side chains, as well as a structure interpreted as being the heavy chain of ITI attached to CS chains. Laser light-scattering revealed that the smaller proteoglycan had a molecular mass of 1.1×106 Da, and EM demonstrated that it had a globular-protein core structure. The core protein, which showed immunological reactivity with perlecan antibodies, was substituted with approximately seven heparan sulphate (HS) and CS chains of similar size (50-55 kDa), the CS disaccharides being mainly 6-sulphated (68%), with a small proportion being 4-sulphated. The protein core was shown to be heterogeneous, with bands occurring at 215, 330 and 400 kDa after enzymic degradation of the glycosaminoglycan chains followed by SDS/PAGE analysis. The demonstration of intact molecules and fragments obtained after stepwise degradations, as shown by gel chromatography, supported a ‘composite’ structure of this proteoglycan.

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