DEVELOPMENT OF RESEARCH REAGENTS SPECIFIC FOR O-GLCNAC (O-GLCNAC LECTENZ)

O-GLCNAC 专用研究试剂 (O-GLCNAC LECTENZ) 的开发

• 批准号: 8744391
• 负责人: LORI YANG
• 金额: $ 100万
• 依托单位: GLYCOSENSORS AND DIAGNOSTICS, LLC
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-13 至 2015-09-12
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8744391
• 关键词: Affinity; Affinity Chromatography; Aliquot; Binding; Biological Assay; Brain; Buffers; Contractor; Custom; Dependence; Detection; Development; Disease; Dissociation; Ensure; Enzyme-Linked Immunosorbent Assay; Enzymes; Equilibrium; Exclusion; Fermentation; Frequencies; Gel; Glycopeptides; Histidine; Immunohistochemistry; Immunoprecipitation; Laboratories; Ligands; Link; Lung; Malignant Neoplasms; Measures; Peptides; Performance; Phase; Polishes; Polymers; Polysaccharides; Printing; Process; Production; Protein p53; Proteins; Protocols documentation; Rattus; Reaction; Reagent; Regulation; Reporting; Reproducibility; Research; Running; Sampling; Services; Silver Staining; Son of Sevenless Proteins; Specificity; Staining method; Stains; Surface Plasmon Resonance; Techniques; Temperature; Testing; Time; Tissues; Universities; Validation; Variant; Western Blotting; base; c-myc Genes; commercialization; design; fast protein liquid chromatography; glycosylation; interest; large scale production; peptide O-linked N-acetylglucosamine-beta-N-acetylglucosaminidase; performance tests; protein aminoacid sequence; prototype; research and development; research study; scale up; screening; tool

The long-term objectives of this proposal are to generate and commercialize a new class of high-specificity, high-affinity proteins called Lectenz¿, as research reagents for use in studies of disease- or cancer-related glycosylation. The specific aims are to further the development and commercialization of a new reagent, created in Phase I of this proposal, which is pan-specific for the detection of the glycan β-O-GlcNAc. By basing the design of this reagent on the biologically-relevant O-GlcNAcase enzyme, the resultant Lectenz¿ should be able to recognize the terminal O-GlcNAc glycan in its biologically-relevant context. For this reason, we refer to this Lectenz¿ as being pan-specific for O-GlcNAc. In this Phase II proposal, we will fully characterize this unique reagent and optimize its production and utility in a variety of assay formats. The ability of the pan-specific O-GlcNAc Lectenz¿ to rapidly confirm the presence of β-O-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 β-O-GlcNAc regulation and associated disease states, such as cancer. In addition, as reported in Phase I, we have found that some clones of this reagent display variations in affinity for O-GlcNAcylated peptides, as a function of the peptide sequence. Thus, we will also continue to select for variants of the Lectenz¿ that may be used to discriminate between β-O-GlcNAcylated peptides, as a function of the peptide sequence.

该提案的长期目标是生成和商业化新的高特异性类别， 高亲和力蛋白称为Lectenz¿，作为用于疾病或癌症相关研究的研究试剂 糖基化。具体目的是进一步开发和商业化新试剂的开发和商业化， 在该提案的第一阶段创建，该提案是用于检测聚糖β-O-GLCNAC的泛特异性。经过 基于该试剂的设计在与生物学上的O-Glcnacase酶上，结果是由生物学 应该能够在与生物学相关的环境中识别末端O-GlcNAC聚糖。为此原因， 我们将此Lectenz®称为O-GLCNAC的泛特异性。在此第二阶段提案中，我们将完全 表征这种独特的试剂，并以各种测定格式优化其生产和实用性。 PAN特异性O-GlcNACLectenz®快速确认蛋白质中存在β-O-GLCNAC的能力 和组织无需转向更精致的技术，将为划定有力的工具 差异化糖基化蛋白质，并最终在调节β-O-GLCNAC和 相关疾病状态，例如癌症。 此外，正如第一阶段报道的那样，我们发现该试剂的一些克隆显示了亲和力的变化 对于O-Glcnacylated Petides，作为肽序列的函数。那，我们还将继续选择 可以用来区分β-O-GlcNacylated肽的Lectenz知识的变体作为功能 胡椒序列。