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Development of a suite of high-specificity fucose binding reagents

一套高特异性岩藻糖结合试剂的开发

基本信息

批准号：
10084298
负责人：
Loretta Yang
金额：
$ 22.5万
依托单位：
GLYCOSENSORS AND DIAGNOSTICS, LLC
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-03-01 至 2023-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10084298
关键词：
Abnormal Cell Active Sites Address Affect Affinity Affinity Chromatography Amino Acids Antibodies Antigens Autoimmune Diseases Binding Biological Markers Carbohydrate Sequence Carbohydrates Cell surface Cells Chemicals Complex Computer Analysis Data Detection Development Directed Molecular Evolution Disease Disease Marker Engineering Enzymes Epitopes Free Energy Fucose Fucosidase Glycoconjugates Glycoproteins Glycoside Hydrolases Human Immune Immune system Immunologic Surveillance In Situ Individual Interferometry Lectin Libraries Ligands Link Location Malignant Neoplasms Malignant neoplasm of pancreas Malignant neoplasm of prostate Mass Spectrum Analysis Measurement Metabolism Methods Monosaccharides Mutagenesis Pancreas Phase Plant Lectins Polymers Polysaccharides Positioning Attribute Preparation Proliferating Property Prostate Protein Subunits Proteins Reagent Recombinants Rheumatoid Arthritis Sampling Site Site-Directed Mutagenesis Specificity Spectrometry Structure Substrate Specificity Surface Techniques Thermotoga maritima Tissue Sample Toxic effect Western Blotting Yeasts arm base cancer type carbohydrate binding protein design differential expression genetic linkage analysis genetic manipulation glycosylation glycosyltransferase histological stains improved in vivo in vivo imaging insight molecular dynamics mutant novel preference response scaffold screening

项目摘要

PROJECT SUMMARY Glycans have several distinct properties that make them excellent targets for disease biomarkers. Firstly, their location glycans on cell surfaces makes them the first point of contact for cellular interactions, and thus they are crucial in the control of normal metabolic processes. Secondly, cell surface molecules are also strategically exposed for surveillance by the immune system allowing for the potential of immune recognition of abnormal cells. Thirdly, specific glycan structures that are not present, or are in low amounts, in normal states proliferate in disease states, such as cancer. And lastly, changes in glycosylation involve many proteins, including those that are highly abundant. Therefore, a single change in a cell’s glycosylation machinery can affect many different glycoconjugates. To effectively employ and discover glycan disease markers a wide range of highly-specific reagents are urgently needed. The monosaccharide fucose has been identified in many disease markers including pancreatic and prostate cancer, but is difficult to detect specifically with existing reagents. Using structurally-guided directed evolution, we will convert an α-L-fucosidase enzyme into high affinity reagents for the detection of fucose-containing antigens. Such engineered lectin-like reagents derived from enzymes are called “Lectenz®”, and have several advantages over lectins and antibodies. The advantages of Lectenz® include precise definition of specificity, tunable binding properties, and ease of recombinant expression, enabling their potential use in affinity purification, western blotting, in situ histological staining, and in vivo imaging. We will generate fucose-binding Lectenz® that can differentiate between various antigenic fucose structures, such as the Lewis antigens versus core fucosylation in found in mammalian glycosylation. Glycosylation detection is essential in fully characterizing and exploiting glycans as markers of specific disease states, and yet current reagents have poor abilities to discriminate between only Lewis X or Lewis A antigens, or other Lewis antigens or core fucose. The principle advantages of an engineered Lectenz® over an antibody are that the Lectenz® is specific to the carbohydrate sequence, but, in contrast to antibodies, will recognize that sequence in a broad range of glycans. Further, in contrast to carbohydrate reagents based on plant lectins, engineered Lectenz® are derived from enzymes that have exquisite substrate specificities and low toxicities.

项目总结