Isolation and characterization of proteoglycans from human follicular fluid
Eriksen, Gitte ; Carlstedt, Ingmar ; Mörgelin, Matthias LU ; Uldbjerg, N and Malmström, Anders LU
(1999)
In Biochemical Journal
340.
p.613-620
- Abstract
- Two proteoglycans differing in size and composition were isolated from human follicular fluid. The larger one of high density had a molecular mass of 3.0x10(6) 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-alpha-trypsin inhibitor (ITI). Western blotting of the CS proteoglycan showed that this had reactivity with antibodies... (More)
- Two proteoglycans differing in size and composition were isolated from human follicular fluid. The larger one of high density had a molecular mass of 3.0x10(6) 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-alpha-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. 1x10(6) 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. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/1115478
- author
- Eriksen, Gitte
; Carlstedt, Ingmar
; Mörgelin, Matthias
LU
; Uldbjerg, N
and Malmström, Anders
LU
- organization
- publishing date
- 1999
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- chondroitin sulphate, heparan sulphate, inter-a-trypsin inhibitor (ITI), proteoglycan, versican-like
- in
- Biochemical Journal
- volume
- 340
- pages
- 613 - 620
- publisher
- Portland Press
- external identifiers
-
- pmid:10359644
- scopus:0033564938
- ISSN
- 0264-6021
- language
- English
- LU publication?
- yes
- id
- 58527f17-4120-4ec2-8b59-8ba37330fd91 (old id 1115478)
- alternative location
- http://www.biochemj.org/bj/340/0613/bj3400613.htm
- date added to LUP
- 2016-04-01 16:38:33
- date last changed
- 2025-10-14 10:13:34
@article{58527f17-4120-4ec2-8b59-8ba37330fd91,
abstract = {{Two proteoglycans differing in size and composition were isolated from human follicular fluid. The larger one of high density had a molecular mass of 3.0x10(6) 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-alpha-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. 1x10(6) 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.}},
author = {{Eriksen, Gitte and Carlstedt, Ingmar and Mörgelin, Matthias and Uldbjerg, N and Malmström, Anders}},
issn = {{0264-6021}},
keywords = {{chondroitin sulphate; heparan sulphate; inter-a-trypsin inhibitor (ITI); proteoglycan; versican-like}},
language = {{eng}},
pages = {{613--620}},
publisher = {{Portland Press}},
series = {{Biochemical Journal}},
title = {{Isolation and characterization of proteoglycans from human follicular fluid}},
url = {{http://www.biochemj.org/bj/340/0613/bj3400613.htm}},
volume = {{340}},
year = {{1999}},
}