Abstract
A glial hyaluronate-binding protein (GHAP) with an isoelectric point of 4.3-4.4 was isolated from human brain white matter. The 60-kDa glycoprotein appeared to be quite resistant to proteolysis, and comparison with GHAP from a viable glioma removed at surgery showed that the protein isolated from autopsy material was not a degradation product resulting from postmortem autolysis. The protein was localized immunohistochemically with mouse monoclonal and rabbit polyclonal antibodies in cerebral white matter. Only small amounts could be found in the gray matter. After enzymatic deglycosylation, an immunoreactive 47-kDa polypeptide was obtained. Two amino acid sequences of GHAP showed a striking similarity (up to 89%) with a highly conserved region of cartilage proteins (bovine nasal cartilage proteoglycan and rat and chicken link protein). However, the amino acid composition and other amino acid sequences suggested that there are also differences between brain-specific GHAP and cartilage proteins.
Highlights
Zymatic deglycosylation, an immunoreactive 47-kDa No immunoreactive material was detected outside the brain polypeptide was obtained
In SDS, urea, or guanidine HC1 extracts of white matter, only one polypeptide was detected on immunoblots corresponding to the 62-kDA band, but after extraction with 10 mM HCl, the molecular mass of the protein in most preparations was reduced to 60 kDa
We report on the isolation and partial characterization of GHAP, a 60-kDa glycoprotein derived from humanbrain white matter
Summary
GHAP was isolated as described under"Materials and Methods." The yield of the preparation was high. In some preparations and depending on theduration of the homogenization of the dissected material inthe HC1 solution, one but two bands, 62 and 60 kDa, were observed Both polypeptides were immunoreactive and appeared to have the same isoelectric point. SDS and HC1 extracts were immunoblotted with the antibodies which were preincubated overnight with purified GHAP. Both the monoclonal and polyclonal antibodies losttheir ability to recognize a bcd FIG. The incubation mixtures were used for immunoblot on HCl extract of human white matte(rlane I ) and purified GHAP (lane2 ) as described under “Materials and Methods.” a, immunoblot with RR50; b, immunoblot with RR50 preincubated with GHAP; c, immunoblot with the rabbit antiserumd; , immunoblot with the rabbit antiserum prein- FIG.. Immunodetection on acid extracts and purified GHAP from spinal cord showed a highdegree of proteolytic degradation
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
