Molecular Modeling and Affinity Determination of scFv Antibody: Proper Linker Peptide Enhances Its Activity

One of existing strategies to engineer active antibody is to link VH and VL domains via a linker peptide. How the composition, length, and conformation of the linker affect antibody activity, however, remains poorly understood. In this study, a dual approach that coordinates molecule modeling, biological measurements, and affinity evaluation was developed to quantify the binding activity of a novel stable miniaturized anti-CD20 antibody or singlechain fragment variable (scFv) with a linker peptide. Upon computer-guided homology modeling, distance geometry analysis, and molecular superimposition and optimization, three new linker peptides PT1, PT2, and PT3 with respective 7, 10, and 15 residues were proposed and three engineered antibodies were then constructed by linking the cloned VH and VL domains and fusing to a derivative of human IgG1. The binding stability and activity of scFv-Fc chimera to CD20 antigen was quantified using a micropipette adhesion frequency assay and a Scatchard analysis. Our data indicated that the binding affinity was similar for the chimera with PT2 or PT3 and ~24-fold higher than that for the chimera with PT1, supporting theoretical predictions in molecular modeling. These results further the understanding in the impact of linker peptide on antibody structure and activity.

生物制剂对沙地樟子松苗木成活生长及生理特征的影响

采用Pt菌剂、ABT生根粉、HRC吸水剂和GGR 6植物生长调节剂等生物制剂,研究了不同生物制剂对2年生樟子松苗木田间试验的造林成活率、植株高度、单株生物量、细根量和盆栽试验苗木的叶片光合和水分利用效率等生理特征的影响。结果表明:(1)Pt3(3号菌剂)、ABT(ABT生根粉)和HRC(HRC吸水剂)均显著地提高樟子松苗木的成活率,它们分别比对照处理苗木成活率高29.3%、23.6%和9.4%,而GRR(GGR植物生长调节剂)和Pt1(1号菌剂)处理苗木的造林成活率同对照处理苗木成活率差异不显著;(2)Pt3、ABT处理均显著地促进了苗木细根生长,分别比对照处理多34.72和15.18 cm,而HRC、CGR和Pt1同对照处理没有显著差异;回归分析表明细根量与苗木成活率呈显著正相关,说明生物制剂提高成活率是通过细根量的增加来实现的;(3)Pt3和ABT处理的苗木高度比对照多2.2、1.5cm和1.1cm,单株生物量干重分别大1.73、0.72g和0.33g,针叶长度、单个针叶面积和单株植株针叶数量分别多1.54和0.096cm、0.188cm2和0.116cm2、31和23束;而Pt1和GGR处理对苗木高度和单株生物量影响不显著;(4)Pt3和ABT处理显著地提高了苗木叶片叶绿素含量和光合速率及水分利用效率等生理指标方面,而其它生物制剂对苗木生理指标影响不显著。生物制剂通过提高苗木生长量和增强生理活动提高樟子松沙地造林成活率。