COVALENT SURFACE IMMOBILIZATION OF ARG-GLY-ASP-CONTAINING AND TYR-ILE-GLY-SER-ARG-CONTAINING PEPTIDES TO OBTAIN WELL-DEFINED CELL-ADHESIVE SUBSTRATES
COVALENT SURFACE IMMOBILIZATION OF ARG-GLY-ASP-CONTAINING AND TYR-ILE-GLY-SER-ARG-CONTAINING PEPTIDES TO OBTAIN WELL-DEFINED CELL-ADHESIVE SUBSTRATES
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DOI：
10.1016/0003-2697(90)90459-m
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发表时间：
1990-06-01
影响因子：
2.9
通讯作者：
HUBBELL, JA
中科院分区：
文献类型：
--
作者：
MASSIA, SP;HUBBELL, JA
合成了含有两类重要细胞粘附受体配体Arg-Gly-Asp（RGD）和Tyr-Ile-Gly-Ser-Arg（YIGSR）的合成肽Gly-Arg-Gly-Asp-Tyr和Gly-Tyr-Ile-Gly-Ser-Arg-Tyr。N-末端Gly作为间隔区，C-末端Y作为放射性碘标记位点。这些改性的基板支持培养的人包皮成纤维细胞（HFFs）的粘附和扩散的独立吸附的蛋白质，它被证明，共价固定的YIGSR含有肽具有生物活性。通过125 I放射性标记测量玻璃上接枝肽的表面浓度，其为12.1 pmol/cm 2。HFF在两种固定化肽底物上扩散，但在接枝YIGSR玻璃表面上的速率比在含RGD的底物上慢得多。在存在或不存在血清的情况下，细胞在RGD衍生的底物上形成局灶性接触。只有当血清存在于培养基中时，YIGSR接枝表面上才形成焦点接触，并且具有与含RGD的基底上观察到的形态不同的形态。血清影响微丝的组织和粘附细胞的扩散程度，虽然粘附蛋白的吸附是最小的所有基板上。这种衍生化方法产生化学稳定的底物，其可用于研究受体介导的细胞粘附，因为可精确测量和控制表面处可用的肽的量。
The synthetic peptides Gly-Arg-Gly-Asp-Tyr and Gly-Tyr-Ile-Gly-Ser-Arg-Tyr, which contain Arg-Gly-Asp (RGD) and Tyr-Ile-Gly-Ser-Arg (YIGSR), the ligands for two important classes of cell adhesion receptors, were covalently coupled to a nonadhesive modified glass surface by the N-terminal Gly. The N-terminal Gly served as a spacer, and the C-terminal Y served as a site for radioiodination. These modified substrates supported the adhesion and spreading of cultured human foreskin fibroblasts (HFFs) independently of adsorbed proteins and, it was demonstrated that a covalently immobilized YIGSR-containing peptide has biological activity. The surface concentration of grafted peptide on the glass was measured by 125I radiolabelling and was 12.1 pmol/cm2. HFFs spread on both immobilized peptide substrates, but at much slower rates on grafted YIGSR glass surfaces than on the RGD-containing substrates. Cells formed focal contacts on the RGD-derivatized substrates in the presence or absence of serum. Focal contacts formed on the YIGSR-grafted surfaces only when serum was present in the medium and had morphologies different from those observed on the RGD-containing substrates. Serum influenced the organization of microfilaments and the extent of spreading of adherent cells, although adsorption of adhesion proteins was minimal on all substrates. This derivatization method produced chemically stable substrates which may be useful in studying receptor-mediated cell adhesion, as the quantity of peptide available at the surface may be precisely measured and controlled.