In-depth Site-specific Analysis of N-glycoproteome in Human Cerebrospinal Fluid and Glycosylation Landscape Changes in Alzheimer's Disease
As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-...
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Published in | Molecular & cellular proteomics Vol. 20; p. 100081 |
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Main Authors | , , , , , , , , , , |
Format | Journal Article |
Language | English |
Published |
United States
Elsevier Inc
01.01.2021
American Society for Biochemistry and Molecular Biology |
Subjects | |
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Abstract | As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-specific CSF N-glycoproteome are lacking. Initial efforts into site-specific N-glycoproteomics study in CSF yield limited coverage, hindering further experimental design of glycosylation-based disease biomarker discovery in CSF. In the present study, we have developed an N-glycoproteomic approach that combines enhanced N-glycopeptide sequential enrichment by hydrophilic interaction chromatography (HILIC) and boronic acid enrichment with electron transfer and higher-energy collision dissociation (EThcD) for large-scale intact N-glycopeptide analysis. The application of the developed approach to the analyses of human CSF samples enabled identifications of a total of 2893 intact N-glycopeptides from 511 N-glycosites and 285 N-glycoproteins. To our knowledge, this is the largest site-specific N-glycoproteome dataset reported for CSF to date. Such dataset provides molecular basis for a better understanding of the structure–function relationships of glycoproteins and their roles in CNS-related physiological and pathological processes. As accumulating evidence suggests that defects in glycosylation are involved in Alzheimer's disease (AD) pathogenesis, in the present study, a comparative in-depth N-glycoproteomic analysis was conducted for CSF samples from healthy control and AD patients, which yielded a comparable N-glycoproteome coverage but a distinct expression pattern for different categories of glycoforms, such as decreased fucosylation in AD CSF samples. Altered glycosylation patterns were detected for a number of N-glycoproteins including alpha-1-antichymotrypsin, ephrin-A3 and carnosinase CN1 etc., which serve as potentially interesting targets for further glycosylation-based AD study and may eventually lead to molecular elucidation of the role of glycosylation in AD progression.
[Display omitted]
•Efficient N-glycopeptide sequential enrichment by HILIC and boronic acid enrichment.•Site-specific intact N-glycopeptide characterization using EThcD.•In-depth site-specific N-glycoproteome analysis in human CSF.•Mapping the landscape of glycosylation patterns in Alzheimer's disease.
An exploratory glycosylation-based biomarker study has been conducted for in-depth mapping of an overall glycosylation landscape and site-specific alteration in glycoproteome collected from cerebrospinal fluids (CSF) in healthy control and Alzheimer’s disease (AD) subjects. The comparison will shed light on the glycoproteome profile, dominant glycosylation differences and similarities, and some of the interesting glycoprotein candidates with specific glycosylation pattern alterations in AD. |
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AbstractList | As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-specific CSF N-glycoproteome are lacking. Initial efforts into site-specific N-glycoproteomics study in CSF yield limited coverage, hindering further experimental design of glycosylation-based disease biomarker discovery in CSF. In the present study, we have developed an N-glycoproteomic approach that combines enhanced N-glycopeptide sequential enrichment by hydrophilic interaction chromatography (HILIC) and boronic acid enrichment with electron transfer and higher-energy collision dissociation (EThcD) for large-scale intact N-glycopeptide analysis. The application of the developed approach to the analyses of human CSF samples enabled identifications of a total of 2893 intact N-glycopeptides from 511 N-glycosites and 285 N-glycoproteins. To our knowledge, this is the largest site-specific N-glycoproteome dataset reported for CSF to date. Such dataset provides molecular basis for a better understanding of the structure–function relationships of glycoproteins and their roles in CNS-related physiological and pathological processes. As accumulating evidence suggests that defects in glycosylation are involved in Alzheimer's disease (AD) pathogenesis, in the present study, a comparative in-depth N-glycoproteomic analysis was conducted for CSF samples from healthy control and AD patients, which yielded a comparable N-glycoproteome coverage but a distinct expression pattern for different categories of glycoforms, such as decreased fucosylation in AD CSF samples. Altered glycosylation patterns were detected for a number of N-glycoproteins including alpha-1-antichymotrypsin, ephrin-A3 and carnosinase CN1 etc., which serve as potentially interesting targets for further glycosylation-based AD study and may eventually lead to molecular elucidation of the role of glycosylation in AD progression.
•
Efficient N-glycopeptide sequential enrichment by HILIC and boronic acid enrichment.
•
Site-specific intact N-glycopeptide characterization using EThcD.
•
In-depth site-specific N-glycoproteome analysis in human CSF.
•
Mapping the landscape of glycosylation patterns in Alzheimer's disease.
An exploratory glycosylation-based biomarker study has been conducted for in-depth mapping of an overall glycosylation landscape and site-specific alteration in glycoproteome collected from cerebrospinal fluids (CSF) in healthy control and Alzheimer’s disease (AD) subjects. The comparison will shed light on the glycoproteome profile, dominant glycosylation differences and similarities, and some of the interesting glycoprotein candidates with specific glycosylation pattern alterations in AD. As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-specific CSF N-glycoproteome are lacking. Initial efforts into site-specific N-glycoproteomics study in CSF yield limited coverage, hindering further experimental design of glycosylation-based disease biomarker discovery in CSF. In the present study, we have developed an N-glycoproteomic approach that combines enhanced N-glycopeptide sequential enrichment by hydrophilic interaction chromatography (HILIC) and boronic acid enrichment with electron transfer and higher-energy collision dissociation (EThcD) for large-scale intact N-glycopeptide analysis. The application of the developed approach to the analyses of human CSF samples enabled identifications of a total of 2893 intact N-glycopeptides from 511 N-glycosites and 285 N-glycoproteins. To our knowledge, this is the largest site-specific N-glycoproteome dataset reported for CSF to date. Such dataset provides molecular basis for a better understanding of the structure-function relationships of glycoproteins and their roles in CNS-related physiological and pathological processes. As accumulating evidence suggests that defects in glycosylation are involved in Alzheimer's disease (AD) pathogenesis, in the present study, a comparative in-depth N-glycoproteomic analysis was conducted for CSF samples from healthy control and AD patients, which yielded a comparable N-glycoproteome coverage but a distinct expression pattern for different categories of glycoforms, such as decreased fucosylation in AD CSF samples. Altered glycosylation patterns were detected for a number of N-glycoproteins including alpha-1-antichymotrypsin, ephrin-A3 and carnosinase CN1 etc., which serve as potentially interesting targets for further glycosylation-based AD study and may eventually lead to molecular elucidation of the role of glycosylation in AD progression.As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-specific CSF N-glycoproteome are lacking. Initial efforts into site-specific N-glycoproteomics study in CSF yield limited coverage, hindering further experimental design of glycosylation-based disease biomarker discovery in CSF. In the present study, we have developed an N-glycoproteomic approach that combines enhanced N-glycopeptide sequential enrichment by hydrophilic interaction chromatography (HILIC) and boronic acid enrichment with electron transfer and higher-energy collision dissociation (EThcD) for large-scale intact N-glycopeptide analysis. The application of the developed approach to the analyses of human CSF samples enabled identifications of a total of 2893 intact N-glycopeptides from 511 N-glycosites and 285 N-glycoproteins. To our knowledge, this is the largest site-specific N-glycoproteome dataset reported for CSF to date. Such dataset provides molecular basis for a better understanding of the structure-function relationships of glycoproteins and their roles in CNS-related physiological and pathological processes. As accumulating evidence suggests that defects in glycosylation are involved in Alzheimer's disease (AD) pathogenesis, in the present study, a comparative in-depth N-glycoproteomic analysis was conducted for CSF samples from healthy control and AD patients, which yielded a comparable N-glycoproteome coverage but a distinct expression pattern for different categories of glycoforms, such as decreased fucosylation in AD CSF samples. Altered glycosylation patterns were detected for a number of N-glycoproteins including alpha-1-antichymotrypsin, ephrin-A3 and carnosinase CN1 etc., which serve as potentially interesting targets for further glycosylation-based AD study and may eventually lead to molecular elucidation of the role of glycosylation in AD progression. As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-specific CSF N-glycoproteome are lacking. Initial efforts into site-specific N-glycoproteomics study in CSF yield limited coverage, hindering further experimental design of glycosylation-based disease biomarker discovery in CSF. In the present study, we have developed an N-glycoproteomic approach that combines enhanced N-glycopeptide sequential enrichment by hydrophilic interaction chromatography (HILIC) and boronic acid enrichment with electron transfer and higher-energy collision dissociation (EThcD) for large-scale intact N-glycopeptide analysis. The application of the developed approach to the analyses of human CSF samples enabled identifications of a total of 2893 intact N-glycopeptides from 511 N-glycosites and 285 N-glycoproteins. To our knowledge, this is the largest site-specific N-glycoproteome dataset reported for CSF to date. Such dataset provides molecular basis for a better understanding of the structure-function relationships of glycoproteins and their roles in CNS-related physiological and pathological processes. As accumulating evidence suggests that defects in glycosylation are involved in Alzheimer's disease (AD) pathogenesis, in the present study, a comparative in-depth N-glycoproteomic analysis was conducted for CSF samples from healthy control and AD patients, which yielded a comparable N-glycoproteome coverage but a distinct expression pattern for different categories of glycoforms, such as decreased fucosylation in AD CSF samples. Altered glycosylation patterns were detected for a number of N-glycoproteins including alpha-1-antichymotrypsin, ephrin-A3 and carnosinase CN1 etc., which serve as potentially interesting targets for further glycosylation-based AD study and may eventually lead to molecular elucidation of the role of glycosylation in AD progression. As the body fluid that directly interchanges with the extracellular fluid of the central nervous system (CNS), cerebrospinal fluid (CSF) serves as a rich source for CNS-related disease biomarker discovery. Extensive proteome profiling has been conducted for CSF, but studies aimed at unraveling site-specific CSF N-glycoproteome are lacking. Initial efforts into site-specific N-glycoproteomics study in CSF yield limited coverage, hindering further experimental design of glycosylation-based disease biomarker discovery in CSF. In the present study, we have developed an N-glycoproteomic approach that combines enhanced N-glycopeptide sequential enrichment by hydrophilic interaction chromatography (HILIC) and boronic acid enrichment with electron transfer and higher-energy collision dissociation (EThcD) for large-scale intact N-glycopeptide analysis. The application of the developed approach to the analyses of human CSF samples enabled identifications of a total of 2893 intact N-glycopeptides from 511 N-glycosites and 285 N-glycoproteins. To our knowledge, this is the largest site-specific N-glycoproteome dataset reported for CSF to date. Such dataset provides molecular basis for a better understanding of the structure–function relationships of glycoproteins and their roles in CNS-related physiological and pathological processes. As accumulating evidence suggests that defects in glycosylation are involved in Alzheimer's disease (AD) pathogenesis, in the present study, a comparative in-depth N-glycoproteomic analysis was conducted for CSF samples from healthy control and AD patients, which yielded a comparable N-glycoproteome coverage but a distinct expression pattern for different categories of glycoforms, such as decreased fucosylation in AD CSF samples. Altered glycosylation patterns were detected for a number of N-glycoproteins including alpha-1-antichymotrypsin, ephrin-A3 and carnosinase CN1 etc., which serve as potentially interesting targets for further glycosylation-based AD study and may eventually lead to molecular elucidation of the role of glycosylation in AD progression. [Display omitted] •Efficient N-glycopeptide sequential enrichment by HILIC and boronic acid enrichment.•Site-specific intact N-glycopeptide characterization using EThcD.•In-depth site-specific N-glycoproteome analysis in human CSF.•Mapping the landscape of glycosylation patterns in Alzheimer's disease. An exploratory glycosylation-based biomarker study has been conducted for in-depth mapping of an overall glycosylation landscape and site-specific alteration in glycoproteome collected from cerebrospinal fluids (CSF) in healthy control and Alzheimer’s disease (AD) subjects. The comparison will shed light on the glycoproteome profile, dominant glycosylation differences and similarities, and some of the interesting glycoprotein candidates with specific glycosylation pattern alterations in AD. |
ArticleNumber | 100081 |
Author | Yu, Qing Carlsson, Cynthia Huang, Junfeng Chen, Zhengwei Okonkwo, Ozioma Yu, Qinying Asthana, Sanjay Zhong, Xiaofang Johnson, Jillian Li, Lingjun Shipman, Richard |
Author_xml | – sequence: 1 givenname: Zhengwei surname: Chen fullname: Chen, Zhengwei organization: Department of Chemistry, University of Wisconsin, Madison, Wisconsin, USA – sequence: 2 givenname: Qinying surname: Yu fullname: Yu, Qinying organization: School of Pharmacy, University of Wisconsin, Madison, Wisconsin, USA – sequence: 3 givenname: Qing surname: Yu fullname: Yu, Qing organization: School of Pharmacy, University of Wisconsin, Madison, Wisconsin, USA – sequence: 4 givenname: Jillian surname: Johnson fullname: Johnson, Jillian organization: School of Pharmacy, University of Wisconsin, Madison, Wisconsin, USA – sequence: 5 givenname: Richard orcidid: 0000-0003-4731-5563 surname: Shipman fullname: Shipman, Richard organization: Department of Applied Science, University of Wisconsin-Stout, Menomonie, Wisconsin, USA – sequence: 6 givenname: Xiaofang surname: Zhong fullname: Zhong, Xiaofang organization: School of Pharmacy, University of Wisconsin, Madison, Wisconsin, USA – sequence: 7 givenname: Junfeng surname: Huang fullname: Huang, Junfeng organization: School of Pharmacy, University of Wisconsin, Madison, Wisconsin, USA – sequence: 8 givenname: Sanjay surname: Asthana fullname: Asthana, Sanjay organization: School of Medicine and Public Health, University of Wisconsin, Madison, Wisconsin, USA – sequence: 9 givenname: Cynthia surname: Carlsson fullname: Carlsson, Cynthia organization: School of Medicine and Public Health, University of Wisconsin, Madison, Wisconsin, USA – sequence: 10 givenname: Ozioma surname: Okonkwo fullname: Okonkwo, Ozioma organization: School of Medicine and Public Health, University of Wisconsin, Madison, Wisconsin, USA – sequence: 11 givenname: Lingjun surname: Li fullname: Li, Lingjun email: lingjun.li@wisc.edu organization: Department of Chemistry, University of Wisconsin, Madison, Wisconsin, USA |
BackLink | https://www.ncbi.nlm.nih.gov/pubmed/33862227$$D View this record in MEDLINE/PubMed |
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Keywords | APP ABC site-specific intact glycopeptide characterization AD SDS DMSO DTT IAA PTM glycopeptide enrichment CNS ADRC EThcD PBA TFA N-glycoproteome analysis WGA RCA electron-transfer higher-energy collision induced dissociation (EThcD) FDR cerebrospinal fluid CSF HILIC ConA |
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Title | In-depth Site-specific Analysis of N-glycoproteome in Human Cerebrospinal Fluid and Glycosylation Landscape Changes in Alzheimer's Disease |
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