Heterogeneity of the chondroitin sulfate portion of phosphacan/6B4 proteoglycan regulates its binding affinity for pleiotrophin/heparin binding growth-associated molecule
Heterogeneity of the chondroitin sulfate portion of phosphacan/6B4 proteoglycan regulates its binding affinity for pleiotrophin/heparin binding growth-associated molecule
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DOI：
10.1074/jbc.m305530200
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发表时间：
2003-09-12
影响因子：
4.8
通讯作者：
Yabe, T
中科院分区：
文献类型：
--
作者：
Maeda, N;He, J;Yabe, T
PTPzeta是一种受体类型的蛋白酪氨酸磷酸酶，以硫酸软骨素蛋白多糖的形式合成，以多营养素为配体。该受体的硫酸软骨素部分是与多效营养素高亲和力结合所必需的。在这里，我们从出生后7天(P7)和P12大鼠大脑皮层(分别为Pg-P7和Pg-P12)和P20大鼠全脑(Pg-P20)中纯化了与PTPzeta的胞外结构域相对应的磷酸聚糖。这些制剂的硫酸软骨素在免疫和成分上表现出不同的结构。特别是，只有PG-P20能与识别含有Glca(2S)β1-3GalNAc(6S)二糖单元(D单元)的硫酸软骨素的单抗MO-225反应。对软骨素酶消化产物的分析表明，这些制剂中的主要成分是GlcAbeta1-3GalNAc(4S)二糖单位(A单位)，PG-P20含有1.3%的D单位，而PG-P7和PG-P12中没有检测到D单位。利用表面等离子体共振生物传感器的相互作用分析表明，PG-P20对多营养素的亲和力(解离常数(K-D)=0.14 nM)接近于PG-P7和PG-P12的5倍，尽管所有这些制剂经软骨素酶ABC消化后都表现出与多营养素的低亲和力结合(K-D=1.4，类似于1.6 nM)。我们还发现，鲨鱼软骨硫酸软骨素D含有类似20%的D单元，与多营养素具有中等亲和力(K-D=2.7 nM)，而鲸鱼软骨硫酸软骨素A不与该生长因子结合。这些结果表明，硫酸软骨素的变化在脑内信号转导的调节中起重要作用。
PTPzeta is a receptor-type protein-tyrosine phosphatase that is synthesized as a chondroitin sulfate proteoglycan and uses pleiotrophin as a ligand. The chondroitin sulfate portion of this receptor is essential for high affinity binding to pleiotrophin. Here, we purified phosphacan, which corresponds to the extracellular domain of PTPzeta, from postnatal day 7 (P7) and P12 rat cerebral cortex (PG-P7 and PG-P12, respectively) and from P20 rat whole brain (PG-P20). The chondroitin sulfate of these preparations displayed immunologically and compositionally different structures. In particular, only PG-P20 reacted with the monoclonal antibody MO-225, which recognizes chondroitin sulfate containing the GlcA(2S)beta1-3GalNAc(6S) disaccharide unit (D unit). Analysis of the chondroitinase digestion products revealed that GlcAbeta1-3GalNAc(4S) disaccharide unit (A unit) was the major component in these preparations and that PG-P20 contained 1.3% D unit, which was not detected in PG-P7 and PG-P12. Interaction analysis using a surface plasmon resonance biosensor indicated that PG-P20 had similar to5-fold stronger affinity for pleiotrophin (dissociation constant (K-D) = 0.14 nM) than PG-P7 and PG-P12, although all these preparations showed similar low affinity binding to pleiotrophin after chondroitinase ABC digestion (K-D = 1.4 similar to 1.6 nM). We also found that shark cartilage chondroitin sulfate D containing similar to20% D unit bound to pleiotrophin with moderate affinity (K-D = 2.7 nM), whereas whale cartilage chondroitin sulfate A showed no binding to this growth factor. These results suggest that variation of chondroitin sulfate plays important roles in the regulation of signal transduction in the brain.