ISSN:
0006-3525
Keywords:
Chemistry
;
Polymer and Materials Science
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Chemistry and Pharmacology
Notes:
Laser light-scattering has been used to investigate the size of native proteoglycan aggregates (PGA-aA1) from day-8 chick limb-bud chondrocyte cultures isolated under associative extraction and purification conditions in 0.4M guanidinium chloride (GdnHCl) solution. Dynamic light-scattering measurements yielded a hydrodynamic radius, Rs, of 244 ± 10 nm for PGA-aA1 in 0.4M GdnHCl, and a weight-average molecular weight (Mw) of 150 ± 50 × 106 was obtained from a Zimm plot. Disaggregation in 4.0M GdnHCl aqueous solution yielded proteoglycan subunits (PGS) with Rs = 39 ± 2 nm, Mw = 1.6 ± 0.3 × 106, which reassembled in 0.4M GdnHCl to form “reconstituted native” aggregates (PGA-raA1) with Rs = 121 ± 6 nm, Mw = 17 ± 3 × 106. A second specimen of PGA-aA1 had Rs = 192 ± 10 nm, Mw = 100 ± 10 × 106. The latter value was estimated from an empirical relationship between Mw and Rs. After dissociation, this specimen reassembled to form PGA-raA1 with Rs = 85 ± 5 nm, Mw = 12 ± 1 × 106. These data are compared with those for a specimen of reconstituted aggregate (PGA-A1) that had been extracted under dissociative conditions and then reaggregated by dialysis to 0.4M GdnHCl aqueous solution, for which Rs = 138 ± 9 nm, Mw = 45 ± 8 × 106. From these values, we have calculated the weight-average number of subunits per aggregate Nw: 111 for PGA-aA1 and 12 for raA1 (70 and 7 for the second PGA-aA1 and PGA-raA1 specimen, respectively) as compared to 32 for PGA-A1. The numbers of subunits per aggregate were also determined from electron micrographs of spread specimens. The latter results show the same trends as those obtained by light scattering, but lead in each case to lower numbers of subunits per aggregate. These data demonstrate conclusively that PGA samples exhibit a higher degree of aggregation in solution than visualized in typical electron microscopy (EM) preparations, probably due to disaggregation during EM specimen preparation. Since Nw determined both by light scattering (LS) and by EM are larger for native versus reconstituted aggregate samples, our data point to a more compact aggregation of subunits along the hyaluronic acid (HA) chains in the former.
Additional Material:
7 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/bip.360250512
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