당과학이 영양의 이해를 도와준다

( Carlito B. Lebrilla ) 생화학분자생물학회(구 한국생화학분자생물학회) 2012 생화학분자생물학회 소식 Vol.32 No.2

Invited Mini Review : Analysis and role of oligosaccharides in milk

( L. Renee Ruhaak ),( Carlito B. Lebrilla ) 생화학분자생물학회(구 한국생화학분자생물학회) 2012 BMB Reports Vol.45 No.8

Milk is an important fluid in glycobiology because it contains a number of short carbohydrate chains either free or as glycoconjugates. These compounds as a class are the most abundant component and benefit the infant by developing and maintaining the infant`s gut flora. New and emerging methods for oligosaccharide analysis have been developed to study milk. These methods allow for the rapid profiling of oligosaccharide mixtures with quantitation. With these tools, the role of oligosaccharide in milk is being understood. They further point to how oligosaccharide analysis can be performed, which until now has been very difficult and have lagged significantly those of other biopolymers. [BMB Reports 2012; 45(8): 442-451]

Combined High-Density Lipoprotein Proteomic and Glycomic Profiles in Patients at Risk for Coronary Artery Disease

Krishnan, Sridevi,Huang, Jincui,Lee, Hyeyoung,Guerrero, Andres,Berglund, Lars,Anuurad, Erdembileg,Lebrilla, Carlito B.,Zivkovic, Angela M. American Chemical Society 2015 Journal of proteome research Vol.14 No.12

<P><B>Objectives:</B> To test whether recently developed methods for comprehensive profiling of the high-density lipoprotein (HDL) glycome combined with the HDL proteome can distinguish individuals with coronary artery disease (CAD) from those without. <B>Methods:</B> Twenty subjects at risk for CAD, who underwent diagnostic coronary arteriography, were analyzed. Ten subjects had CAD, and ten did not. HDL was extracted from fasting plasma samples by ultracentrifugation, followed by shotgun proteomic, glycomic, and ganglioside analyses using LC-MS. CAD vs non-CAD subjects’ data were compared using univariate and multivariate statistics. <B>Results:</B> Principal components analysis showed a clear separation of CAD and non-CAD subjects, confirming that combined HDL proteomic and glycomic profiles distinguished at-risk subjects with atherosclerosis from those without. CAD patients had lower HDL apolipoprotein content (specifically ApoA-I, A-II, and E, <I>p</I> < 0.05), and lower serum amyloid A2 (SAA2, p = 0.020) and SAA4 (p = 0.007) but higher sialylated glycans (<I>p</I> < 0.05). <B>Conclusion:</B> Combined proteomic and glycomic profiling of isolated HDL was tested as a novel analytical approach for developing biomarkers of disease. In this pilot study we found that HDL proteome and glycome distinguished between individuals who had CAD from those who did not within a group of individuals equally at risk for heart disease.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jprobs/2015/jprobs.2015.14.issue-12/acs.jproteome.5b00730/production/images/medium/pr-2015-00730j_0008.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/pr5b00730'>ACS Electronic Supporting Info</A></P>

Isomer-Specific LC/MS and LC/MS/MS Profiling of the Mouse Serum N-Glycome Revealing a Number of Novel Sialylated N-Glycans

Hua, Serenus,Jeong, Ha Neul,Dimapasoc, Lauren M.,Kang, Inae,Han, Chanyoung,Choi, Jong-Soon,Lebrilla, Carlito B.,An, Hyun Joo American Chemical Society 2013 ANALYTICAL CHEMISTRY - Vol.85 No.9

<P>Mice are the premier mammalian models for studies of human physiology and disease, bearing extensive biological similarity to humans with far fewer ethical, economic, or logistic complications. To facilitate glycomic studies based on the mouse model, we comprehensively profiled the mouse serum N-glycome using isomer-specific nano-LC/MS and -LC/MS/MS. N-Glycans were identified by accurate mass MS and structurally elucidated by MS/MS. Porous graphitized carbon nano-LC was able to separate out nearly 300 N-linked glycan compounds (including isomers) from just over 100 distinct N-linked glycan compositions. Additional MS/MS structural analysis was performed on a number of novel N-glycans, revealing the structural characteristics of modifications such as dehydration, O-acetylation, and lactylation. Experimental findings were combined with known glycobiology to generate a theoretical library of all biologically possible mouse serum N-glycan compositions. The library may be used for automated identification of complex mixtures of mouse N-glycans, with possible applications to a wide range of mouse-related research endeavors, including pharmaceutical drug development and biomarker discovery.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/ancham/2013/ancham.2013.85.issue-9/ac400195h/production/images/medium/ac-2013-00195h_0007.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/ac400195h'>ACS Electronic Supporting Info</A></P>

Regulating host-microbe interactions by glycomic landscaping

Dayoung Park,Narine Arabyan,Chatchai Phoomak,Bart C. Weimer,Sopit Wongkham,Carlito B. Lebrilla 한국당과학회 2016 한국당과학회 학술대회 Vol.2016 No.01

Cell surface glycans lie at the interface between the cell and its environment, playing an active role in mediating interactions with foreign substances. Although current lectin-based methods enable detection of select glycan epitopes present on the cell surface, detailed structural information of glycan heterogeneity is needed to better assess their involvement in cell-cell interactions, particularly with pathogens. Mass spectrometry-based glycomics has the potential to provide a single platform to monitor hundreds of native structures simultaneously. This study constitutes a comprehensive and quantitative analysis that defines the sequence of oligosaccharides on human cells that affect the nature of Salmonella Typhimurium infection. We first examined the glycan products remaining on the host cell surface after brief contact with bacteria, which included mainly, desialylated and high mannose glycans. We then describe a glycomic landscaping (glycoscaping) strategy to present predominantly high mannose glycans on host cells to better correlate phenotype with structure. A better understanding of the biological roles of glycosylation is achieved by structure-specific re-engineering towards specified glycosylation patterns. Changes in the glycan landscape were induced on live cells, using a mixture of glycosidases and glycosidase inhibitors. Following treatment, the exact changes were characterized by mass spectrometry. Such glycoscaping, when applied to intestinal epithelial cells, yielded significant increases in membrane enzyme activity, cell permeability, migration, and microbial infection.

Characterization of Novel <i>O</i>-Glycans Isolated from Tear and Saliva of Ocular Rosacea Patients

Ozcan, Sureyya,An, Hyun Joo,Vieira, Ana C.,Park, Gun Wook,Kim, Jae Han,Mannis, Mark J.,Lebrilla, Carlito B. American Chemical Society 2013 JOURNAL OF PROTEOME RESEARCH Vol.12 No.3

<P><I>O</I>-Glycans in saliva and tear isolated from patients suffering from ocular rosacea, a form of inflammatory ocular surface disease, were profiled, and their structures were elucidated using high resolution mass spectrometry. We have previously shown that certain structures, particularly sulfated oligosaccharides, increased in the tear and saliva of rosacea patients. In this study, the structures of these glycans were elucidated using primarily tandem mass spectrometry. There were important similarities in the glycan profiles of tears and saliva with the majority of the structures in common. The structures of the most abundant species common to both tear and saliva, which were also the most abundant species in both, were elucidated. For sulfated species, the positions of the sulfate groups were localized. The majority of the structures were new, with the sulfated glycans comprising mucin <I>core 1</I>- and <I>core 2</I>-type structures. As both saliva and tear are rich in mucins, it is suggested that the <I>O</I>-glycans are mainly components of mucins. The study further illustrates the strong correspondence between the glycans in the tear and saliva of ocular rosacea patients.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jprobs/2013/jprobs.2013.12.issue-3/pr3008013/production/images/medium/pr-2012-008013_0010.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/pr3008013'>ACS Electronic Supporting Info</A></P>

Isomer-specific chromatographic profiling yields highly sensitive and specific potential N-glycan biomarkers for epithelial ovarian cancer

Hua, S.,Williams, C.C.,Dimapasoc, L.M.,Ro, G.S.,Ozcan, S.,Miyamoto, S.,Lebrilla, C.B.,An, H.J.,Leiserowitz, G.S. Elsevier 2013 Journal of chromatography A Vol.1279 No.-

Aberrant glycosylation has been observed for decades in essentially all types of cancer, and is now well established as an indicator of carcinogenesis. Mining the glycome for biomarkers, however, requires analytical methods that can rapidly separate, identify, and quantify isomeric glycans. We have developed a rapid-throughput method for chromatographic glycan profiling using microfluidic chip-based nanoflow liquid chromatography (nano-LC)/mass spectrometry. To demonstrate the utility of this method, we analyzed and compared serum samples from epithelial ovarian cancer cases (n=46) and healthy control individuals (n=48). Over 250 N-linked glycan compound peaks with over 100 distinct N-linked glycan compositions were identified. Statistical testing identified 26 potential glycan biomarkers based on both compositional and structure-specific analyses. Using these results, an optimized model was created incorporating the combined abundances of seven potential glycan biomarkers. The receiver operating characteristic (ROC) curve of this optimized model had an area under the curve (AUC) of 0.96, indicating robust discrimination between cancer cases and healthy controls. Rapid-throughput chromatographic glycan profiling was found to be an effective platform for structure-specific biomarker discovery.

Extensive Determination of Glycan Heterogeneity Reveals an Unusual Abundance of High Mannose Glycans in Enriched Plasma Membranes of Human Embryonic Stem Cells

An, Hyun Joo,Gip, Phung,Kim, Jaehan,Wu, Shuai,Park, Kun Wook,McVaugh, Cheryl T.,Schaffer, David V.,Bertozzi, Carolyn R.,Lebrilla, Carlito B. American Society for Biochemistry and Molecular Bi 2012 Molecular and Cellular Proteomics Vol.11 No.4

Glyco-Analytical Multispecific Proteolysis (Glyco-AMP): A Simple Method for Detailed and Quantitative Glycoproteomic Characterization

Hua, Serenus,Hu, Chloe Y.,Kim, Bum Jin,Totten, Sarah M.,Oh, Myung Jin,Yun, Nayoung,Nwosu, Charles C.,Yoo, Jong Shin,Lebrilla, Carlito B.,An, Hyun Joo American Chemical Society 2013 Journal of proteome research Vol.12 No.10

<P>Despite recent advances, site-specific profiling of protein glycosylation remains a significant analytical challenge for conventional proteomic methodology. To alleviate the issue, we propose glyco-analytical multispecific proteolysis (Glyco-AMP) as a strategy for glycoproteomic characterization. Glyco-AMP consists of rapid, in-solution digestion of an analyte glycoprotein (or glycoprotein mixture) by a multispecific protease (or protease cocktail). Resulting glycopeptides are chromatographically separated by isomer-specific porous graphitized carbon nano-LC, quantified by high-resolution MS, and structurally elucidated by MS/MS. To demonstrate the consistency and customizability of Glyco-AMP methodology, the glyco-analytical performances of multispecific proteases subtilisin, pronase, and proteinase K were characterized in terms of quantitative accuracy, sensitivity, and digestion kinetics. Glyco-AMP was shown be effective on glycoprotein mixtures as well as glycoproteins with multiple glycosylation sites, providing detailed, quantitative, site- and structure-specific information about protein glycosylation.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jprobs/2013/jprobs.2013.12.issue-10/pr400442y/production/images/medium/pr-2013-00442y_0010.gif'></P>

Identification and Accurate Quantitation of Biological Oligosaccharide Mixtures

Strum, John S.,Kim, Jaehan,Wu, Shuai,De Leoz, Maria Lorna A.,Peacock, Kyle,Grimm, Rudolf,German, J. Bruce,Mills, David A.,Lebrilla, Carlito B. American Chemical Society 2012 ANALYTICAL CHEMISTRY - Vol.84 No.18

<P>Structure-specific characterization and quantitation is often required for effective functional studies of oligosaccharides. Inside the gut, HMOs are preferentially bound and catabolized by the beneficial bacteria. HMO utility by these bacteria employs structure-specific catabolism based on a number of glycosidases. Determining the activity of these enzymes requires accurate quantitation of a large number of structures. In this study, we describe a method for the quantitation of human milk oligosaccharide (HMO) structures employing LC/MS and isotopically labeled internal standards. Data analysis was accomplished with a newly developed software tool, LC/MS Searcher, that employs a reference structure library to process LC/MS data yielding structural identification with accurate quantitation. The method was used to obtain a meta-enzyme analysis of bacteria, the simultaneous characterization of all glycosidases employed by bacteria for the catabolism of milk oligosaccharides. Analysis of consumed HMO structures confirmed the utility of a β-1,3-galactosidase in <I>Bifidobacterium longum subsp. infantis</I> ATCC 15697 (<I>B. infantis</I>). In comparison, <I>Bifidobacterium breve</I> ATCC 15700 showed significantly less HMO catabolic activity compared to <I>B. infantis</I>.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/ancham/2012/ancham.2012.84.issue-18/ac301128s/production/images/medium/ac-2012-01128s_0002.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/ac301128s'>ACS Electronic Supporting Info</A></P>