SBIR TOPIC 294 PHASE 1 - DEVELOPMENT OF GLYCOSYLATION - SPECIFIC RESEARCH

SBIR 主题 294 第 1 阶段 - 糖基化的发展 - 特定研究

• 批准号: 8354002
• 负责人: LORI YANG
• 金额: $ 14.89万
• 依托单位: GLYCOSENSORS AND DIAGNOSTICS, LLC
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-15 至 2012-06-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8354002
• 关键词: Affinity; Antibodies; Antigen Targeting; Antigens; Binding; Biosensor; Biotin; Chromatography; Computing Methodologies; Detection; Development; Disease; Enzymes; Equilibrium; Escherichia coli; Exclusion; Glycopeptides; Glycoproteins; Homologous Gene; Human; Ion Exchange; Libraries; Malignant Neoplasms; Measures; Methods; Monosaccharides; Mutagenesis; Peptides; Phage Display; Phase; Polysaccharides; Preparation; Probability; Production; Proteins; Reagent; Regulation; Reporting; Research; Screening procedure; Site; Site-Directed Mutagenesis; Small Business Innovation Research Grant; Specificity; Surface Plasmon Resonance; System; Techniques; Testing; Tissues; Toxic effect; Variant; aptamer; base; c-myc Genes; cost; design; fast protein liquid chromatography; glycosylation; in vivo; innovation; instrument; protein aminoacid sequence; success; tool; vector

The long term objectives of this proposal are to demonstrate the utility of a new class of high-specificity high-affinity proteins called Lectenz¿ as research reagents for use in recognition of disease- or cancer-related glycosylation sites. The specific aims are to create a reagent that is pan-specific for the detection of the glycan f3-0-GlcNAc itself, as well as variations of this reagent that are able to detect f3-0-GIcNAc in the context of its disease- or cancer-specific peptide sequence (glycosylation site). The ability to rapidly confirm the presence of f3-0-GlcNAc in proteins and tissues without the need to turn to more-elaborate techniques would provide a powerful tool to delineate differentially glycosylated proteins and eventually establish correlations between f3-0-GIcNAc regulation and associated disease states, such as cancer. Using a combination of point mutagenesis, display library screening, and forefront computational methods, the human enzyme O-GIcNAcase (hOGA) will be converted into a high-specificity biosensor for its natural substrate f3-0-GIcNAc. Lectenz¿ have several potential advantages over antibodies or aptamers, including predefined specificity for the target antigen, ease of preparation in a monovalent form, and (for human homologs) a low probability of in vivo toxicity. The innovative use of computational methods results in efficiency gains in the design and screening of the display library, lowering costs and increasing success rates.

该提案的长期目标是证明一类名为 Lectenz 的新型高特异性高亲和力蛋白作为研究试剂用于识别疾病或癌症相关糖基化位点的效用。具体目标是创建一种泛特异性试剂，用于检测聚糖 f3-0-GlcNAc 本身，以及该试剂的变体，能够在其疾病或癌症特异性肽序列（糖基化位点）背景下检测 f3-0-GlcNAc。无需求助于更精细的技术即可快速确认蛋白质和组织中 f3-0-GlcNAc 的存在，这将为描述差异糖基化蛋白质提供强大的工具，并最终建立 f3-0-GlcNAc 调节与相关疾病状态（例如癌症）之间的相关性。通过结合点诱变、展示库筛选和前沿计算方法，人类酶 O-GlcNAcase (hOGA) 将被转化为其天然底物 f3-0-GlcNAc 的高特异性生物传感器。与抗体或适体相比，Lectenz 具有几个潜在的优势，包括对目标抗原的预定义特异性、易于以单价形式制备，以及（对于人类同系物）体内毒性的可能性较低。计算方法的创新使用提高了设计和筛选的效率 展示库，降低成本并提高成功率。