Proteases and other components involved in the modification of chloroplast structures:
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
1998
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| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | 133 S. Ill., graph. Darst. |
Internformat
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Datensatz im Suchindex
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| adam_text | 6 PROTEASES AND OTHER COMPONENTS INVOLVED IN THE MODIFICATION OF
CHLOROPLAST STRUCTURES INAUGURAL-DISSERTATION ZUR ERIANGUNG DES
DOKTORGRADES DER FAKULTAT FUR BIOLOGIE DER
LUDWIG-MAXIMILIANS-UNIVERSITST MONCHEN VORGELEGTVON ANNA SOKOLENKO AUS
ODESSA, UKRAINE 1998 CONTENT 1 CONTENT ABBREVIATIONS 5 1. INTRODUCTION 8
2. MATERIALS AND METHODS 16 2.1 MATERIALS 16 2.1.1 CHEMICALS AND ENZYMES
16 2.1.2 BACTERIAL STRAINS 16 2.1.3 VECTORS 16 2.1.4
RECOMBINANTPLASMIDS(CDNA INSERTS) 17 2.1.5 PLANT MATERIAL 17 2.1.6
MOLECULAR WEIGHT STANDARDS 17 2.1.6.1 DNA AND RNA MOLECULAR WEIGHT
STANDARDS 17 2.1.6.2 PROTEIN MOLECULAR WEIGHT STANDARDS 18 2.1.7 PRIMERS
18 2.1.8 MEDIA FOR GROWTH OF BACTERIA AND PHAGES 19 2.1.9 GENERAL
BUFFERS AND SOLUTIONS 20 2.1.10 ANTIBODIES 20 2.2 METHODS 20 2.2.1
STANDARD METHODS 20 2.2.2 ISOLATION AND ANALYSIS OF NUCLEIC ACIDS FROM
PLANTS 21 2.2.2.1 ISOLATION OF GENOMIC DNA FROM PLANTS 21 2.2.2.2
ISOLATION OF POLY(A) + RNA USING OLIGO(DT)-CELLULOSE 22 2.2.2.3
RADIOACTIVE LABELLING OF DNA FRAGMENTS WITH KLENOW ENZYME .... 23
2.2.2.4 HYBRIDIZATION WITH RADIBLABELLED DNA FRAGMENTS 24 2.2.3 USE
OFRECOMBINANTPHAGE DNA LIBRARIES 24 2.2.3.1 SCREENING OF PHAGE CDNA
LIBRARIES 24 2.2.3.2 ISOLATION OF PHAGE DNA 25 2.2.4 GEL ELECTROPHORESIS
OF PROTEINS 26 2.2.4.1 SDS-DENATURING PAA GELS 26 2.2.4.2 NON-DENATURING
GEL ELECTROPHORESIS FOR SOLUBLE PROTEIN FRACTIONS ... 26 2.2.4.3
NON-DENATURING GEL ELECTROPHORESIS FOR MEMBRANE PROTEIN FRACTIONS 27
2.2.4.4 TWO-DIMENSIONAL GEL ELCTROPHORESIS 28 2.2.4.5 COOMASSIE
BRILLIANT BLUE STAINING OF PAA GELS 28 2.2.4.6 SILVER STAINING OF PAA
GELS 29 2.2.5 ANALYSIS OF PROTEINS ON NITROCELLULOSE MEMBRANES 29
CONTENT 2.2.5.1 TRANSFER OF SDS-DENATURED PROTEINS ONTO NITROCELLULOSE
MEMBRANES 29 2.2.5.2 STAINING BLOTS WITH PONCEAU S 30 2.2.6 ISOLATION
AND FRACTIONATION OF CHLOROPLASTS 30 2.2.6.1 ISOLATION OF INTACT
CHLOROPLASTS 30 2.2.6.2 FRACTIONATION OF CHLOROPLASTS INTO STROMA,
THYLAKOID MEMBRANES AND LUMENAL COMPONENTS 31 2.2.6.2.1 ISOLATION OF
THYLAKOID MEMBRANES 31 2.2.6.2.2 ISOLATION OF LUMENAL PROTEINS 32 2.2.7
DETERMINATION OF PROTEIN CONCENTRATIONS 33 2.2.8 TRANSCRIPTION IN VITRO
34 2.2.9 TRANSLATION IN VITRO 35 2.2.10 IN ORGANELLO IMPORT EXPERIMENTS
36 2.2.11 OVEREXPRESSION OF PROTEINS IN VITRO AND PURIFICATION OF
OVEREXPRESSED PROTEINS 37 2.2.11.1 CLONING INTO OVEREXPRESSION VECTORS
37 2.2.11.2 OVEREXPRESSION OF PROTEINS IN E. COLI 38 2.2.11.3 PARTIAL
PURIFICATION OF OVEREXPRESSED PLASTOCYANIN 38 2.2.12 SEPARATION OF CLP
COMPLEXES BY SIZE EXCLUSION CHROMATOGRAPHY 40 2.2.13 ANTIBODY GENERATION
AND USE 40 2.2.13.1 ANTIBODY GENERATION 40 2.2.13.2 PURIFICATION OF
ANTIBODIES AGAINST CONTAMINATIONS WITH E. COLI PROTEINS 41 2.2.13.3
IMMUNOLOGICAL DETECTION OF PROTEINS ON MEMBRANES 42 2.2.13.3.1 WESTERN
ANALYSIS USING ALKALINE PHOSPHATASE-CONJUGATED ANTIBODIES 42 2.2.13.3.2
WESTERN ANALYSIS USING HORSERADISH PEROXIDASE-CONJUGATED ANTIBODIES 42
2.2.13.4 IMMUNOPRECIPITATION OF THE PROTEINS WITH POLYCLONAL ANTISERA 43
2.2.14 ASSAYS FOR PROTEOLYTIC ACTIVITIES 43 2.2.14.1 DETECTION OF
PROTEOLYTIC ACTIVITIES BY MEASURING THE RELEASED AMINO ACIDS 43 2.2.14.2
SUBSTRATE-CONTAINING DENATURING GELS 44 2.2.14.3 DETECTION OF PROTEASE
ACTIVITIES USING AZOCOLL 44 2.2.14.4 PARTIAL PURIFICATION OF SDS-STABLE
PROTEASES ON GELATIN-AGAROSE COLUMNS . 44 2.2.15 ASSAY FOR INORGANIC
PYROPHOSPHATASE ACTIVITY 45 CONTENT 3. RESULTS 46 3.1 PROTEOLYSIS IN
CHLOROPLASTS 46 3.1.1 DEGRADATION PROCESSES IN CHLOROPLASTS 46 3.1.1.1
CHARACTERIZATION OF THE DEGRADATION PROCESSES IN CHLOROPLAST
SUBCOMPARTMENTS 46 3.1.1.1.1 VISUALIZATION OF PROTEOLYTIC ACTIVITIES IN
CHLOROPLAST SUBCOMPARTMENTS 46 3.1.1.1.2 PROTEOLYTIC ACTIVITIES IN
THYLAKOID MEMBRANES 47 3.1.1.1.3 SUBSTRATE SPECIFICITY OF THYLAKOID
PROTEASES 48 3.1.2 DEGRADATION OF PLASTOCYANIN 49 3.1.2.1 DEPENDENCE OF
PLASTOCYANIN DEGRADATION ON THE SUBSTRATE 49 3.1.2.2 PH DEPENDENCE OF
PLASTOCYANIN DEGRADATION 54 3.1.3 SDS-STABLE PROTEASES 55 3.1.3.1
OPTIMIZATION OF PROTEASE IDENTIFICATION ON SUBSTRATE-CONTAINING GELS *.
55 3.1.3.2 IDENTIFICATION OF SDS-STABLE PROTEASES ON
SUBSTRATE-CONTAINING GELS 56 3.1.3.3 BIOCHEMICAL CHARACTERIZATION 59
3.1.3.4 PREPARATIVE ISOLATION OF SDS-STABLE PROTEASES 60 3.1.4 CLP
PROTEASES 62 3.1.4.1 THE CATALYTIC SUBUNIT CLPC 62 3.1.4.1.1 ISOLATION
OF CLPC CDNA 62 3.1.4.1.2 TRANSLOCATIONOFCLPC INTO THE CHLOROPLASTS 67
3.1.4.2 THE PROTEOLYTIC SUBUNIT CLPP 68 3.1.4.2.1 NUCLEAR-ENCODED CLPP
68 3.1.4.2.1.1 N-CLPP CDNA 68 3.1.4.2.1.2 TRANSLOCATION OF NCLPP INTO
CHLOROPLASTS AND PREDICTION OF PROCESSING SITES 70 3.1.4.2.2
PLASTID-ENCODEDCLPP 73 3.1.4.2.2.1 P-CLPP DNA AND CONSTRUCTION OF
CIMNENC PCLPP .... 73 3.1.4.2.2.2 IMPORT OF CHIMERIC PCLPP IN ORGANE/ZO
73 -3.1.4.3 SUPRAMOLECULAR CLP COMPLEXES 74 CONTENT 3.2 MINUTE
COMPONENTS OF THYLAKOID MEMBRANES 79 3.2.1 IDENTIFICATION OFNEW
CHLOROPLAST COMPONENTS 79 3.2.2 CHARACTERIZATION OF NEW PROTEINS FROM
THYLAKOID MEMBRANES 79 3.2.2.1 ISOLATION AND CHARACTERIZATION OF TML2 79
3.2.2.1.1 ANALYSIS OF TML2 CDNA 79 3.2.2.1.2 TRANSLOCATION OF TML2 IN IN
ORGANELLO ASSAYS 81 3.2.2.1.3 OVEREXPRESSION OF TML2 PROTEIN IN E. COLI
84 3.2.2.1.4 SOUTHERN, DEVELOPMENTAL NORTHERN AND WESTERN ANALYSIS .. 85
3.2.2.1.5 FUNCTIONAL AND STRUCTURAL ANALYSIS OF TML2 86 3.2.2.1.6
LOCALIZATION OF TML2 WITHIN THYLAKOID MEMBRANES 88 3.2.3 ISOLATION AND
CHARACTERIZATION OF TML4 89 3.2.4 ISOLATION OF POLYPHENOL OXIDASE CDNA
93 3.2.4.1 CHARACTERIZATION OF POLYPHENOL OXIDASE CDNA 93 3.2.4.2 IMPORT
OF PRE-PPO INTO ISOLATED CHLOROPLASTS 96 4. DISCUSSION 97 4.1
DEGRADATION PROCESSES IN CHLOROPLASTS 97 4.1.1 ASSAYS FOR IDENTIFICATION
OF THE CHLOROPLAST PROTEASES 97 4.1.2 LOCATION OF PROTEASES IN
CHLOROPLASTS 99 4.1.3 SDS-STABLE PROTEASES 100 4.1.4 CLPPROTEASES 101
4.1.4.1 COMPLEX FORMATION OF CLP PROTEASES 101 4.1.4.2 EVOLUTION OF
CLPPS 104 4.2 CHARACTERIZATION OF NOVEL THYLAKOID COMPONENTS 110 5.
REFERENCES 1 14 6. SUMMARY 126 7. APPENDIX 129 7.1 CLPP GENES FROM
DIFFERENT ORGANISMS 129 ACKNOWLEDGMENTS 13 1 PUBLICATIONS 132 CURRICULUM
VITAE 133
|
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| indexdate | 2024-07-09T18:24:28Z |
| institution | BVB |
| language | English |
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| physical | 133 S. Ill., graph. Darst. |
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| spelling | Sokolenko, Anna 1970- Verfasser (DE-588)120841487 aut Proteases and other components involved in the modification of chloroplast structures von Anna Sokolenko 1998 133 S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier München, Univ., Diss., 1998 Chloroplast (DE-588)4147821-6 gnd rswk-swf Struktur (DE-588)4058125-1 gnd rswk-swf Proteasen (DE-588)4047521-9 gnd rswk-swf Modifikation Biologie (DE-588)4268640-4 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Chloroplast (DE-588)4147821-6 s Struktur (DE-588)4058125-1 s Modifikation Biologie (DE-588)4268640-4 s Proteasen (DE-588)4047521-9 s DE-604 GBV Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008308944&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Sokolenko, Anna 1970- Proteases and other components involved in the modification of chloroplast structures Chloroplast (DE-588)4147821-6 gnd Struktur (DE-588)4058125-1 gnd Proteasen (DE-588)4047521-9 gnd Modifikation Biologie (DE-588)4268640-4 gnd |
| subject_GND | (DE-588)4147821-6 (DE-588)4058125-1 (DE-588)4047521-9 (DE-588)4268640-4 (DE-588)4113937-9 |
| title | Proteases and other components involved in the modification of chloroplast structures |
| title_auth | Proteases and other components involved in the modification of chloroplast structures |
| title_exact_search | Proteases and other components involved in the modification of chloroplast structures |
| title_full | Proteases and other components involved in the modification of chloroplast structures von Anna Sokolenko |
| title_fullStr | Proteases and other components involved in the modification of chloroplast structures von Anna Sokolenko |
| title_full_unstemmed | Proteases and other components involved in the modification of chloroplast structures von Anna Sokolenko |
| title_short | Proteases and other components involved in the modification of chloroplast structures |
| title_sort | proteases and other components involved in the modification of chloroplast structures |
| topic | Chloroplast (DE-588)4147821-6 gnd Struktur (DE-588)4058125-1 gnd Proteasen (DE-588)4047521-9 gnd Modifikation Biologie (DE-588)4268640-4 gnd |
| topic_facet | Chloroplast Struktur Proteasen Modifikation Biologie Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008308944&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT sokolenkoanna proteasesandothercomponentsinvolvedinthemodificationofchloroplaststructures |