The Resource Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins, Sudheer Bobba
Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins, Sudheer Bobba
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Resource Information
The item Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins, Sudheer Bobba represents a specific, individual, material embodiment of a distinct intellectual or artistic creation found in Missouri University of Science & Technology Library.
This item is available to borrow from all library branches.
- Summary
- The research presented in this dissertation was aimed at understanding several structural and biochemical features underlying the bacterial cell wall biosynthetic process using statistical, biochemical and analytical tools. Penicillin-binding proteins (PBPs) are ubiquitous and essential enzymes necessary for cell wall biosynthesis. PBPs are broadly classified as low molecular mass (LMM) and high molecular mass (HMM) based on their molecular weight. Although all PBPs share a set of highly conserved active site residues, different PBPs have different propensities for catalyzing reactions. To understand the structural differences among PBPs, a global catalytic residue comparison was performed for all the PBPs available in the protein data bank. This dataset was analyzed using univariate and multivariate statistical methods, revealing several interesting relationships such as: (1) Distribution of the dihedral angle for the SXXK-motif Lys side chain was bimodal, and strongly correlated with HMM/transpeptidase vs LMM/hydrolase activity; (2) Distance between the SXXK-motif Lys-NZ atom and the Lys/His-nitrogen atom of the (K/H)T(S)G-motif was highly conserved. Unlike LMM PBPs, HMM PBPs when purified, give undetectable or weak enzyme activity that has impeded efforts to develop new inhibitors. In the next section of this dissertation, we described a fluorescently detected microtiter plate-based assay for HMM PBPs. Purified PBPs were immobilized onto microtiter plate wells and labeled with biotinylated-ampicillin (Bi--Amp). Treatment of Bi--Amp-labeled PBPs with a streptavidin--horseradish peroxidase conjugate followed by a fluorogenic HRP substrate allowed the detection of PBPs. The HMM PBP assay was extended to penicillin-binding protein 2a, the molecular determinant of high-level [beta]-lactam resistance in methicillin-resistant Staphylococcus aureus using Bio-Amp and biotinylated-cephalexin as tagging reagents. This assay was then demonstrated for use in competition assays for screening and characterizing both [beta]-lactam and non [beta]-lactam inhibitors. The cytoplasmic steps of cell wall biosynthesis involve a series of UDP-linked intermediates whose synthesis is catalyzed by Mur enzymes. The final section of the dissertation deals with developing an LC-MS/MS detection and quantification of UDP-peptidoglycan intermediates in the bacterial cell wall biosynthetic pathway. The assay was validated and then used to quantitate the in vivo levels of UDP-intermediates in S. aureus treated with different antibiotics acting at various stages of cell wall biosynthesis
- Language
- eng
- Extent
- 1 online resource (266 pages)
- Note
-
- "A dissertation in Pharmaceutical Sciences and Chemistry."
- Advisor: William G. Gutheil
- Vita
- Contents
-
- Introduction and literature review
- Multivariate geometrical analysis of catalytic residues in the penicillin-binding proteins
- Substrate specificity studies of low molecular mass PBP actinomadura R39
- A microtiter plate-based [beta]-lactam binding assay for inhibitors of high-molecular-mass penicillin-binding proteins
- A microtiter plate-based assay for inhibitors of penicillin-binding protein 2A from methicillin-resistant staphylococcus aureus
- Quantitative analysis of cytoplasmic UDP-peptidoglycan intermediates in s. aureus using LC-ESI-MS/MS
- Liquid chromatography-tandem mass spectrometry assay for detection and quantitation of endogenous bioactive peptides in rat brain
- Summary and conclusions
- Appendix
- Label
- Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins
- Title
- Structural and enzymatic characterization of bacterial cell wall enzymes
- Title remainder
- focus on penicillin-binding proteins
- Statement of responsibility
- Sudheer Bobba
- Language
- eng
- Summary
- The research presented in this dissertation was aimed at understanding several structural and biochemical features underlying the bacterial cell wall biosynthetic process using statistical, biochemical and analytical tools. Penicillin-binding proteins (PBPs) are ubiquitous and essential enzymes necessary for cell wall biosynthesis. PBPs are broadly classified as low molecular mass (LMM) and high molecular mass (HMM) based on their molecular weight. Although all PBPs share a set of highly conserved active site residues, different PBPs have different propensities for catalyzing reactions. To understand the structural differences among PBPs, a global catalytic residue comparison was performed for all the PBPs available in the protein data bank. This dataset was analyzed using univariate and multivariate statistical methods, revealing several interesting relationships such as: (1) Distribution of the dihedral angle for the SXXK-motif Lys side chain was bimodal, and strongly correlated with HMM/transpeptidase vs LMM/hydrolase activity; (2) Distance between the SXXK-motif Lys-NZ atom and the Lys/His-nitrogen atom of the (K/H)T(S)G-motif was highly conserved. Unlike LMM PBPs, HMM PBPs when purified, give undetectable or weak enzyme activity that has impeded efforts to develop new inhibitors. In the next section of this dissertation, we described a fluorescently detected microtiter plate-based assay for HMM PBPs. Purified PBPs were immobilized onto microtiter plate wells and labeled with biotinylated-ampicillin (Bi--Amp). Treatment of Bi--Amp-labeled PBPs with a streptavidin--horseradish peroxidase conjugate followed by a fluorogenic HRP substrate allowed the detection of PBPs. The HMM PBP assay was extended to penicillin-binding protein 2a, the molecular determinant of high-level [beta]-lactam resistance in methicillin-resistant Staphylococcus aureus using Bio-Amp and biotinylated-cephalexin as tagging reagents. This assay was then demonstrated for use in competition assays for screening and characterizing both [beta]-lactam and non [beta]-lactam inhibitors. The cytoplasmic steps of cell wall biosynthesis involve a series of UDP-linked intermediates whose synthesis is catalyzed by Mur enzymes. The final section of the dissertation deals with developing an LC-MS/MS detection and quantification of UDP-peptidoglycan intermediates in the bacterial cell wall biosynthetic pathway. The assay was validated and then used to quantitate the in vivo levels of UDP-intermediates in S. aureus treated with different antibiotics acting at various stages of cell wall biosynthesis
- Cataloging source
- UMK
- http://library.link/vocab/creatorDate
- 1986-
- http://library.link/vocab/creatorName
- Bobba, Sudheer
- Degree
- Ph. D. (School of Pharmacy and Dept. of Chemistry).
- Dissertation year
- 2013.
- Granting institution
- University of Missouri-Kansas City,
- Illustrations
- illustrations
- Index
- no index present
- Literary form
- non fiction
- Nature of contents
-
- dictionaries
- bibliography
- theses
- http://library.link/vocab/relatedWorkOrContributorName
- Gutheil, William G.
- http://library.link/vocab/subjectName
-
- Penicillin
- Bacterial cell walls
- Label
- Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins, Sudheer Bobba
- Note
-
- "A dissertation in Pharmaceutical Sciences and Chemistry."
- Advisor: William G. Gutheil
- Vita
- Antecedent source
- not applicable
- Bibliography note
- Includes bibliographical references (pages 213-244)
- Carrier category
- online resource
- Carrier category code
-
- cr
- Carrier MARC source
- rdacarrier.
- Color
- black and white
- Content category
- text
- Content type code
-
- txt
- Content type MARC source
- rdacontent.
- Contents
- Introduction and literature review -- Multivariate geometrical analysis of catalytic residues in the penicillin-binding proteins -- Substrate specificity studies of low molecular mass PBP actinomadura R39 -- A microtiter plate-based [beta]-lactam binding assay for inhibitors of high-molecular-mass penicillin-binding proteins -- A microtiter plate-based assay for inhibitors of penicillin-binding protein 2A from methicillin-resistant staphylococcus aureus -- Quantitative analysis of cytoplasmic UDP-peptidoglycan intermediates in s. aureus using LC-ESI-MS/MS -- Liquid chromatography-tandem mass spectrometry assay for detection and quantitation of endogenous bioactive peptides in rat brain -- Summary and conclusions -- Appendix
- Control code
- 857794172
- Dimensions
- unknown
- Extent
- 1 online resource (266 pages)
- File format
- one file format
- Form of item
- online
- Level of compression
- mixed
- Media category
- computer
- Media MARC source
- rdamedia.
- Media type code
-
- c
- Other physical details
- illustrations (some color)
- Quality assurance targets
- not applicable
- Specific material designation
- remote
- System control number
- (OCoLC)857794172
- System details
-
- The full text of the dissertation is available as an Adobe Acrobat .pdf file; Adobe Acrobat Reader required to view the file
- Mode of access: World Wide Web
- Label
- Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins, Sudheer Bobba
- Note
-
- "A dissertation in Pharmaceutical Sciences and Chemistry."
- Advisor: William G. Gutheil
- Vita
- Antecedent source
- not applicable
- Bibliography note
- Includes bibliographical references (pages 213-244)
- Carrier category
- online resource
- Carrier category code
-
- cr
- Carrier MARC source
- rdacarrier.
- Color
- black and white
- Content category
- text
- Content type code
-
- txt
- Content type MARC source
- rdacontent.
- Contents
- Introduction and literature review -- Multivariate geometrical analysis of catalytic residues in the penicillin-binding proteins -- Substrate specificity studies of low molecular mass PBP actinomadura R39 -- A microtiter plate-based [beta]-lactam binding assay for inhibitors of high-molecular-mass penicillin-binding proteins -- A microtiter plate-based assay for inhibitors of penicillin-binding protein 2A from methicillin-resistant staphylococcus aureus -- Quantitative analysis of cytoplasmic UDP-peptidoglycan intermediates in s. aureus using LC-ESI-MS/MS -- Liquid chromatography-tandem mass spectrometry assay for detection and quantitation of endogenous bioactive peptides in rat brain -- Summary and conclusions -- Appendix
- Control code
- 857794172
- Dimensions
- unknown
- Extent
- 1 online resource (266 pages)
- File format
- one file format
- Form of item
- online
- Level of compression
- mixed
- Media category
- computer
- Media MARC source
- rdamedia.
- Media type code
-
- c
- Other physical details
- illustrations (some color)
- Quality assurance targets
- not applicable
- Specific material designation
- remote
- System control number
- (OCoLC)857794172
- System details
-
- The full text of the dissertation is available as an Adobe Acrobat .pdf file; Adobe Acrobat Reader required to view the file
- Mode of access: World Wide Web
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<div class="citation" vocab="http://schema.org/"><i class="fa fa-external-link-square fa-fw"></i> Data from <span resource="http://link.library.mst.edu/portal/Structural-and-enzymatic-characterization-of/u1-jk3V5vM8/" typeof="Book http://bibfra.me/vocab/lite/Item"><span property="name http://bibfra.me/vocab/lite/label"><a href="http://link.library.mst.edu/portal/Structural-and-enzymatic-characterization-of/u1-jk3V5vM8/">Structural and enzymatic characterization of bacterial cell wall enzymes : focus on penicillin-binding proteins, Sudheer Bobba</a></span> - <span property="potentialAction" typeOf="OrganizeAction"><span property="agent" typeof="LibrarySystem http://library.link/vocab/LibrarySystem" resource="http://link.library.mst.edu/"><span property="name http://bibfra.me/vocab/lite/label"><a property="url" href="http://link.library.mst.edu/">Missouri University of Science & Technology Library</a></span></span></span></span></div>
