Biglycan isoforms with differences in polysaccharide substitution and core protein in human lung fibroblasts.
Biglycan is widely distributed in the extracellular matrix and is a member of the small proteoglycan family characterized by a core protein with leucine-rich repeat motifs. We show in this paper for the first time that biglycan from human lung fibroblasts can be expressed as different isoforms. These isoforms can be separated from the predominant form of biglycan by hydrophobic interaction chromatography, where the more hydrophobic isoforms are retarded. The newly found isoforms of biglycan have a smaller core protein substituted with smaller glycosaminoglycan chains, migrating on SDS/PAGE at between 110 and 200 kDa. These molecules were identified as biglycan using MALDI-TOF MS. Identification of C-terminal peptides together with glycosylation of the N-terminal glycosaminoglycan sites excludes the possibility of terminal proteolytic cleavage. The biglycan isoforms are N-glycosylated, which demonstrates that a lack in N-glycosylation is not the reason for a smaller core. Two components revealed by RT-PCR indicate alternative splicing, which could be located in regions of the protein that have not been identified, with the exclusion of sites of glycosylations. Analyses of glycosaminoglycan chain length of the isoforms show that besides the normally occurring glycosaminoglycan chains, there is a mixture of shorter glycosaminoglycan chains. Structural analysis shows that these glycosaminoglycan chains contain a lower proportion of iduronic acid (61%) relative to glucuronic acid when compared to the glycosaminoglycan chain of the predominant form of biglycan (71%). We can anticipate that variation in structure of biglycan can cause changes in the connective tissue formation depending on its ability to bind matrix molecules, as well as cytokines.
Tufvesson
Ellen
E
Department of Cell and Molecular Biology, BMC, Lund University, Sweden. Ellen.Tufvesson@medkem.lu.se
Malmström
Johan
J
Marko-Varga
György
G
Westergren-Thorsson
Gunilla
G
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D058578:Biglycan; D002478:Cells, Cultured; D002850:Chromatography, Gel / Q000379:methods; D004187:Disaccharides / Q000032:analysis; D016326:Extracellular Matrix Proteins; D005347:Fibroblasts / Q000378:metabolism; D005960:Glucosides / Q000032:analysis; D006031:Glycosylation; D006801:Humans; D007067:Iduronic Acid / Q000032:analysis; D008969:Molecular Sequence Data; D020033:Protein Isoforms; D011509:Proteoglycans / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
C546884:BGN protein, human; D058578:Biglycan; D004187:Disaccharides; D016326:Extracellular Matrix Proteins; D005960:Glucosides; D020033:Protein Isoforms; D011509:Proteoglycans; C447240:geranyl 6-O-alpha-L-arabinopyranosyl-beta-D-glucopyranoside; D007067:Iduronic Acid