公开(公告)号：US07332613B2

公开(公告)日：2008-02-19

申请号：US10863589

申请日：2004-06-07

申请人： Steven P. Gygi ,  Scott Anthony Gerber ,  Carlos Augusto Gartner

发明人： Steven P. Gygi ,  Scott Anthony Gerber ,  Carlos Augusto Gartner

IPC分类号： C07D235/02

CPC分类号： G01N33/6848 ,  G01N33/6842 ,  G01N2458/15 ,  Y10T436/24

摘要： The invention provides non-affinity based isotope tagged peptides, chemistries for making these peptides, and methods for using these peptides. In one aspect, tags comprise a reactive site (RS) for reacting with a molecule on a protein to form a stable association with the peptide (e.g., a covalent bond) and an anchoring site (AS) group for reversibly or removably anchoring the tag to a solid phase such as a resin support. Anchoring may be direct or indirect (e.g., through a linker molecule). Preferably, the anchoring site comprises a biotin compound. Preferably, the tag comprises a mass-altering label, such as a stable isotope, such that association of the tag with the peptide can be monitored by mass spectrometry. The reagents can be used for rapid and quantitative analysis of proteins or protein function in mixtures of proteins.

摘要翻译： 本发明提供了基于非亲和力的同位素标记的肽，用于制备这些肽的化学物质和使用这些肽的方法。 一方面，标签包含用于与蛋白质上的分子反应以与肽（例如共价键）和锚定位点（AS）组稳定结合的反应位点（RS），用于可逆地或可拆卸地锚定标签 固相如树脂载体。 锚定可以是直接的或间接的（例如通过连接体分子）。 优选地，锚定位点包含生物素化合物。 优选地，标签包含质量变化标签，例如稳定同位素，使得可以通过质谱法监测标签与肽的缔合。 这些试剂可用于蛋白质混合物中蛋白质或蛋白质功能的快速和定量分析。

公开(公告)号：US20090053742A1

公开(公告)日：2009-02-26

申请号：US11985768

申请日：2007-11-16

申请人： Steven P. Gygi ,  Scott Anthony Gerber ,  Carlos Augusto Gartner

发明人： Steven P. Gygi ,  Scott Anthony Gerber ,  Carlos Augusto Gartner

IPC分类号： G01N33/543 ,  G01N33/574

CPC分类号： G01N33/6848 ,  G01N33/6842 ,  G01N2458/15 ,  Y10T436/24

摘要： The invention provides non-affinity based isotope tagged peptides, chemistries for making these peptides, and methods for using these peptides. In one aspect, tags comprise a reactive site (RS) for reacting with a molecule on a protein to form a stable association with the peptide (e.g., a covalent bond) and an anchoring site (AS) group for reversibly or removably anchoring the tag to a solid phase such as a resin support. Anchoring may be direct or indirect (e.g., through a linker molecule). Preferably, the anchoring site comprises a biotin compound. Preferably, the tag comprises a mass-altering label, such as a stable isotope, such that association of the tag with the peptide can be monitored by mass spectrometry. The reagents can be used for rapid and quantitative analysis of proteins or protein function in mixtures of proteins.

摘要翻译： 本发明提供了基于非亲和力的同位素标记的肽，用于制备这些肽的化学物质和使用这些肽的方法。 在一个方面，标签包含用于与蛋白质上的分子反应以与肽（例如共价键）和锚定位点（AS）组稳定结合的反应位点（RS），用于可逆地或可拆卸地锚定标签 固相如树脂载体。 锚定可以是直接的或间接的（例如通过连接体分子）。 优选地，锚定位点包含生物素化合物。 优选地，标签包含质量变化标签，例如稳定同位素，使得可以通过质谱法监测标签与肽的缔合。 这些试剂可用于蛋白质混合物中蛋白质或蛋白质功能的快速和定量分析。

公开(公告)号：US07501286B2

公开(公告)日：2009-03-10

申请号：US10781047

申请日：2004-02-17

申请人： Steven P. Gygi ,  Scott Anthony Gerber

发明人： Steven P. Gygi ,  Scott Anthony Gerber

IPC分类号： G01N24/00

CPC分类号： G01N33/6848 ,  G01N33/68 ,  G01N33/6842 ,  Y10T436/24

摘要： The invention provides reagents, kits and methods for detecting and/or quantifying proteins in complex mixtures, such as a cell lysate. The methods can be used in high throughput assays to profile cellular proteomes. In one aspect, the invention provides a peptide internal standard labeled with a stable isotope and corresponding in amino acid sequence to the amino acid sequence of a subsequence of a target polypeptide. In another aspect, the peptide internal standard is labeled at a modified amino acid residue and is used to determine the presence of, and/or quantitate the amount of a particular modified form of a protein.

摘要翻译： 本发明提供用于检测和/或定量复杂混合物中的蛋白质的试剂，试剂盒和方法，例如细胞裂解物。 该方法可用于高通量测定以分析细胞蛋白质组。 在一个方面，本发明提供了用稳定同位素标记并在氨基酸序列中对应于靶多肽的亚序列的氨基酸序列的肽内标。 在另一方面，肽内标在修饰的氨基酸残基处标记，并用于测定蛋白质的特定修饰形式的量的存在和/或定量。

公开(公告)号：US20130252950A1

公开(公告)日：2013-09-26

申请号：US13825723

申请日：2011-09-23

申请人： John Blenis ,  Steven P. Gygi ,  Yonghao Yu

发明人： John Blenis ,  Steven P. Gygi ,  Yonghao Yu

IPC分类号： C12Q1/48 ,  G01N33/68

CPC分类号： C12Q1/485 ,  G01N33/5011 ,  G01N33/68 ,  G01N33/74 ,  G01N2333/4703

摘要： Provided are over 300 mTOR kinase targets identified by a comprehensive phosphoproteomics assay. Methods of targeting mTOR kinase targets, methods to determine the level of mTOR activity by measuring the level of phosphorylation of an mTOR targeted phosphorylation site, methods for distinguishing different classes of mTOR activity in a cell based on phosphoproteomic analysis of mTOR-targeted proteins are also provided. Also provided is the classification of a hyperproliferative disease based on phosphoproteomic analysis of mTOR-targeted proteins, as well as the personalization of therapeutic methods for the treatment of hyperproliferative disease based on phosphoproteomics. Also provided are therapeutic methods including administering to a subject an mTOR inhibitor, an mTOR inhibitor and an additional kinase inhibitor, or a dual inhibitor of mTOR and an additional kinase based on the phosphorylation levels of mTOR targets determined in the subject. Some aspects of this invention relate to the discovery that GrblO is an mTOR-targeted tumor suppressor gene.

摘要翻译： 提供了通过综合磷酸化蛋白质测定法鉴定的超过300个mTOR激酶靶标。 靶向mTOR激酶靶标的方法，通过测量mTOR靶向磷酸化位点的磷酸化水平来测定mTOR活性水平的方法，基于mTOR靶向蛋白质的磷酸化蛋白质分析来区分细胞中不同种类的mTOR活性的方法也是 提供。 还提供了基于mTOR靶向蛋白质的磷酸化蛋白质组学分析的过度增殖性疾病的分类，以及基于磷酸化蛋白质处理过度增殖性疾病的治疗方法的个性化。 还提供了治疗方法，包括向受试者施用mTOR抑制剂，mTOR抑制剂和另外的激酶抑制剂，或mTOR的双重抑制剂和基于受试者确定的mTOR靶标的磷酸化水平的另外的激酶。 本发明的一些方面涉及GrblO是mTOR靶向的肿瘤抑制基因的发现。

公开(公告)号：US06670194B1

公开(公告)日：2003-12-30

申请号：US09383062

申请日：1999-08-25

申请人： Rudolf Hans Aebersold ,  Michael H. Gelb ,  Steven P. Gygi ,  C. Ronald Scott ,  Frantisek Turecek ,  Scott A. Gerber ,  Beate Rist

发明人： Rudolf Hans Aebersold ,  Michael H. Gelb ,  Steven P. Gygi ,  C. Ronald Scott ,  Frantisek Turecek ,  Scott A. Gerber ,  Beate Rist

IPC分类号： C07C33116

CPC分类号： G01N33/6848 ,  C12Q1/25 ,  G01N33/58 ,  G01N33/6803 ,  G01N33/6842 ,  Y10S530/812 ,  Y10T436/182 ,  Y10T436/24 ,  Y10T436/25 ,  Y10T436/25125 ,  Y10T436/25375

摘要： Analytical reagents and mass spectrometry-based methods using these reagents for the rapid, and quantitative analysis of proteins or protein function in mixtures of proteins. The methods employ affinity labeled protein reactive reagents having three portions: an affinity label (A) covalently linked to a protein reactive group (PRG) through a linker group (L). The linker may be differentially isotopically labeled, e.g., by substitution of one or more atoms in the linker with a stable isotope thereof. These reagents allow for the selective isolation of peptide fragments or the products of reaction with a given protein (e.g., products of enzymatic reaction) from complex mixtures. The isolated peptide fragments or reaction products are characteristic of the presence of a protein or the presence of a protein function in those mixtures. Isolated peptides or reaction products are characterized by mass spectrometric (MS) techniques. The reagents also provide for differential isotopic labeling of the isolated peptides or reaction products which facilitates quantitative determination by mass spectrometry of the relative amounts of proteins in different samples. The methods of this invention can be used for qualitative and quantitative analysis of global protein expression profiles in cells and tissues, to screen for and identify proteins whose expression level in cells, tissue or biological fluids is affected by a stimulus or by a change in condition or cell state of the cell, tissue or organism from which the sample originated.

摘要翻译： 使用这些试剂的分析试剂和基于质谱的方法用于蛋白质混合物中蛋白质或蛋白质功能的快速和定量分析。 该方法采用具有三个部分的亲和标记的蛋白质反应试剂：通过连接基团（L）与蛋白质反应基团（PRG）共价连接的亲和标记（A）。 连接体可以是差异同位素标记的，例如用连接体中的一个或多个原子用其稳定同位素取代。 这些试剂允许从复杂混合物中选择性分离肽片段或与给定蛋白质（例如，酶反应产物）反应的产物。 分离的肽片段或反应产物是这些混合物中存在蛋白质或存在蛋白质功能的特征。 分离的肽或反应产物的特征在于质谱（MS）技术。 试剂还提供分离的肽或反应产物的差异同位素标记，其有助于通过质谱法定量测定不同样品中蛋白质的相对量。 本发明的方法可用于细胞和组织中全球蛋白质表达谱的定性和定量分析，筛选和鉴定细胞，组织或生物流体中的表达水平受刺激或病情变化影响的蛋白质 或来自样品的细胞，组织或生物体的细胞状态。

公开(公告)号：US06852544B2

公开(公告)日：2005-02-08

申请号：US09839884

申请日：2001-04-20

申请人： Rudolf Hans Aebersold ,  Michael H. Gelb ,  Steven P. Gygi ,  C. Ronald Scott ,  Frantisek Turecek ,  Scott A. Gerber ,  Beate Rist

发明人： Rudolf Hans Aebersold ,  Michael H. Gelb ,  Steven P. Gygi ,  C. Ronald Scott ,  Frantisek Turecek ,  Scott A. Gerber ,  Beate Rist

IPC分类号： G01N27/62 ,  C12Q1/25 ,  C12Q1/34 ,  C12Q1/48 ,  G01N30/00 ,  G01N30/06 ,  G01N30/60 ,  G01N30/72 ,  G01N30/88 ,  G01N33/53 ,  G01N33/566 ,  G01N33/58 ,  G01N33/68

CPC分类号： G01N33/6848 ,  C12Q1/25 ,  G01N33/58 ,  G01N33/6803 ,  G01N33/6842 ,  Y10S530/812 ,  Y10T436/182 ,  Y10T436/24 ,  Y10T436/25 ,  Y10T436/25125 ,  Y10T436/25375

摘要： Analytical reagents and mass spectrometry-based methods using these reagents for the rapid, and quantitative analysis of proteins or protein function in mixtures of proteins. The methods employ affinity labeled protein reactive reagents having three portions: an affinity label (A) covalently linked to a protein reactive group (PRG) through a linker group (L). The linker may be differentially isotopically labeled, e.g., by substitution of one or more atoms in the linker with a stable isotope thereof. These reagents allow for the selective isolation of peptide fragments or the products of reaction with a given protein (e.g., products of enzymatic reaction) from complex mixtures. The isolated peptide fragments or reaction products are characteristic of the presence of a protein or the presence of a protein function in those mixtures. Isolated peptides or reaction products are characterized by mass spectrometric (MS) techniques. The reagents also provide for differential isotopic labeling of the isolated peptides or reaction products which facilitates quantitative determination by mass spectrometry of the relative amounts of proteins in different samples. The methods of this invention can be used for qualitative and quantitative analysis of global protein expression profiles in cells and tissues, to screen for and identify proteins whose expression level in cells, tissue or biological fluids is affected by a stimulus or by a change in condition or state of the cell, tissue or organism from which the sample originated.

摘要翻译： 使用这些试剂的分析试剂和基于质谱的方法用于蛋白质混合物中蛋白质或蛋白质功能的快速和定量分析。 该方法采用具有三个部分的亲和标记的蛋白质反应试剂：通过连接基团（L）与蛋白质反应基团（PRG）共价连接的亲和标记（A）。 连接体可以是差异同位素标记的，例如用连接体中的一个或多个原子用其稳定同位素取代。 这些试剂允许从复杂混合物中选择性分离肽片段或与给定蛋白质（例如，酶反应产物）反应的产物。 分离的肽片段或反应产物是这些混合物中存在蛋白质或存在蛋白质功能的特征。 分离的肽或反应产物的特征在于质谱（MS）技术。 试剂还提供分离的肽或反应产物的差异同位素标记，其有助于通过质谱法定量测定不同样品中蛋白质的相对量。 本发明的方法可用于细胞和组织中全球蛋白质表达谱的定性和定量分析，筛选和鉴定细胞，组织或生物流体中的表达水平受刺激或病情变化影响的蛋白质 或来自样品的细胞，组织或生物体的状态。

公开(公告)号：US20120149883A1

公开(公告)日：2012-06-14

申请号：US13284255

申请日：2011-10-28

申请人： Steven P. Gygi ,  Junmin Peng

发明人： Steven P. Gygi ,  Junmin Peng

IPC分类号： C07K17/00 ,  C07K16/00

CPC分类号： G01N33/6848 ,  C07K1/22 ,  C07K7/08 ,  C12Q1/37 ,  G01N33/6803 ,  G01N33/6842 ,  G01N33/6896 ,  G01N2440/36 ,  G01N2560/00 ,  Y10T436/24

摘要： The invention provides a method detecting and quantifying proteins by mass spectrophotometric analysis using peptide internal standards and provides a highly sensitive way of detecting protein modifications. In one aspect, the invention provides a method for determining a site of ubiquitination in a polypeptide and for evaluating ubiquitination targets in a population of polypeptides. In this way, a proteome ubiquitination map can be obtained which comprises information relating to the ubiquitination states of a plurality of cellular polypeptides. Maps can be obtained for a variety of different types of cells and cell states. For example, ubiquitination targets in normal and diseased cells can be evaluated. Preferably, the map is stored as data files in a database. Individual ubiquitinated polypeptides identified can be used to generate molecular probes diagnostic of a cell state and/or can serve as targets for agents that modulate one or more cellular processes.

摘要翻译： 本发明提供了通过使用肽内标的质谱分析法检测和定量蛋白质的方法，并提供了检测蛋白质修饰的高度灵敏的方法。 一方面，本发明提供了一种确定多肽泛素化位点的方法，并用于评价多肽群体中的泛素化靶标。 以这种方式，可以获得包含与多个细胞多肽的泛素化状态有关的信息的蛋白质组泛素化图。 可以获得各种不同类型的细胞和细胞状态的图。 例如，可以评估正常和患病细胞中的泛素化靶标。 优选地，地图作为数据文件存储在数据库中。 鉴定的单个泛素化多肽可用于产生诊断细胞状态的分子探针和/或可用作调节一种或多种细胞过程的试剂的靶标。

公开(公告)号：US07544518B2

公开(公告)日：2009-06-09

申请号：US10994815

申请日：2004-11-23

申请人： Rudolf Hans Aebersold ,  Michael H. Gelb ,  Steven P. Gygi ,  C. Ronald Scott ,  Frantisek Turecek ,  Scott A. Gerber ,  Beate Rist

发明人： Rudolf Hans Aebersold ,  Michael H. Gelb ,  Steven P. Gygi ,  C. Ronald Scott ,  Frantisek Turecek ,  Scott A. Gerber ,  Beate Rist

IPC分类号： G01N33/68

CPC分类号： G01N33/6848 ,  C12Q1/25 ,  G01N33/58 ,  G01N33/6803 ,  G01N33/6842 ,  Y10S530/812 ,  Y10T436/182 ,  Y10T436/24 ,  Y10T436/25 ,  Y10T436/25125 ,  Y10T436/25375

摘要： Analytical reagents and mass spectrometry-based methods using these reagents for the rapid, and quantitative analysis of proteins or protein function in mixtures of proteins. The methods employ affinity labeled protein reactive reagents having three portions: an affinity label (A) covalently linked to a protein reactive group (PRG) through a linker group (L). The linker may be differentially isotopically labeled, e.g., by substitution of one or more atoms in the linker with a stable isotope thereof. These reagents allow for the selective isolation of peptide fragments or the products of reaction with a given protein (e.g., products of enzymatic reaction) from complex mixtures. The isolated peptide fragments or reaction products are characteristic of the presence of a protein or the presence of a protein function in those mixtures. Isolated peptides or reaction products are characterized by mass spectrometric (MS) techniques. The reagents also provide for differential isotopic labeling of the isolated peptides or reaction products which facilitates quantitative determination by mass spectrometry of the relative amounts of proteins in different samples. The methods of this invention can be used for qualitative and quantitative analysis of global protein expression profiles in cells and tissues, to screen for and identify proteins whose expression level in cells, tissue or biological fluids is affected by a stimulus or by a change in condition or state of the cell, tissue or organism from which the sample originated.

公开(公告)号：US20080214782A1

公开(公告)日：2008-09-04

申请号：US11980279

申请日：2007-10-30

申请人： Steven P. Gygi ,  Judit Villen ,  Sean Beausoleil

发明人： Steven P. Gygi ,  Judit Villen ,  Sean Beausoleil

IPC分类号： C07K7/08 ,  C07K5/10 ,  C07K7/06 ,  C07K16/18 ,  C40B40/10 ,  C07K19/00 ,  C07K2/00 ,  C07F9/572

CPC分类号： C07K7/08 ,  C07K14/705 ,  C07K14/715

摘要： The present invention relates to phosphopeptide compositions and anti-phosphopeptide antibody compositions. Also provided are methods of identifying phosphorylation sites in phosphorylated peptides and phosphorylation site motifs.

摘要翻译： 本发明涉及磷酸肽组合物和抗磷酸肽抗体组合物。 还提供了鉴定磷酸化肽和磷酸化位点基序中的磷酸化位点的方法。

公开(公告)号：US08669116B2

公开(公告)日：2014-03-11

申请号：US10506877

申请日：2003-03-11

申请人： Steven P. Gygi ,  Junmin Peng

发明人： Steven P. Gygi ,  Junmin Peng

IPC分类号： C12Q1/37 ,  C07K1/22 ,  G01N33/68

CPC分类号： G01N33/6848 ,  C07K1/22 ,  C07K7/08 ,  C12Q1/37 ,  G01N33/6803 ,  G01N33/6842 ,  G01N33/6896 ,  G01N2440/36 ,  G01N2560/00 ,  Y10T436/24

摘要： The invention provides a method detecting and quantifying proteins by mass spectrophotometric analysis using peptide internal standards and provides a highly sensitive way of detecting protein modifications. In one aspect, the invention provides a method for determining a site of ubiquitination in a polypeptide and for evaluating ubiquitination targets in a population of polypeptides. In this way, a proteome ubiquitination map can be obtained which comprises information relating to the ubiquitination states of a plurality of cellular polypeptides. Maps can be obtained for a variety of different types of cells and cell states. For example, ubiquitination targets in normal and diseased cells can be evaluated. Preferably, the map is stored as data files in a database. Individual ubiquitinated polypeptides identified can be used to generate molecular probes diagnostic of a cell state and/or can serve as targets for agents that modulate one or more cellular processes.

摘要翻译： 本发明提供了通过使用肽内标的质谱分析法检测和定量蛋白质的方法，并提供了检测蛋白质修饰的高度灵敏的方法。 一方面，本发明提供了一种确定多肽泛素化位点的方法，并用于评价多肽群体中的泛素化靶标。 以这种方式，可以获得包含与多个细胞多肽的泛素化状态有关的信息的蛋白质组泛素化图。 可以获得各种不同类型的细胞和细胞状态的图。 例如，可以评估正常和患病细胞中的泛素化靶标。 优选地，地图作为数据文件存储在数据库中。 鉴定的单个泛素化多肽可用于产生诊断细胞状态的分子探针和/或可用作调节一种或多种细胞过程的试剂的靶标。