| dc.description.abstract | Abstract
Xanthomonas campestris pv. campestris is a gram-negative bacterium that is phytophathogenic to crucifers such as Brassics and Arabidopsis and causes black rot. The genus Xanthomonas is a diverse and economically important group of bacterial phytopathogens. Structural genomics, which aims to determine the three-dimensional structures of all proteins on a genome wide scale. The genome of Xanthomonas campestris pv. campestris str. ATCC 33913 was sequenced by ONSA/FAPBSP/Brazil group in 2002 and that of Xanthomonas campestris pv. campestris str. 17 by an integrated structural and functional group in Taiwan in 2002 too. We try to understand, in structure terms, its pathogenicity to its host, its capability to produce xanthan gum, its protein secretion pathway, its different gene regulation behavior, and the proteins produced when it is under stress condition. Another important point about this structural genomics projects is to gain novel structural information. In this thesis we aim to identify and characterize the three-dimensional structures of several proteins in the XCC using high-resolution NMR techniques. In this respect, two XCC proteins, XC 975 and XC 2382, were selected for structural studies from screening 18 target proteins by 2D 15N-1HSQC NMR spectra. XC 975 codes for a hypothetical protein with a molecular mass of 9.7 kDa. A BLASTp search with this sequence revealed that most of the homologous sequences are annoted as hypothetical proteins. In COG database, XC 975 belongs to the family COG0271 named stress-induced morphogen. We have assigned nearly complete resonance of the 1H, 15N, 13C nuclei using 2D 15N -1H HSQC and triple resonance experiments including 3D HNCACB、3D CACB (CO)NH、HNCO、3D H(CC-CO)NH-TOCSY、3D(H)C(C-CO)NH-TOCSY etc. The XC 975 secondary structure topologyα β β α α β as calculated by the CSI program TALOS program using 1HA, 13CA, 13CB, 13C’ and 15N secondary chemical shifts and the complementary 3D NOESY-15N-HSQC spectrum data. XC 2382, with a molecular mass of 14.21 kDa, is named apaG because the gene is located in a multifunctional ksgA-apaG-apaH operon. The apaG expression is also tightly linked to apaH, suggesting that both gene products might be involved in the same biological function related to the metabolism of Ap4N. The unusual nucleotides (Ap4N) could be involved in the priming reaction of replication or synthesized as alarmones to signal the outset of cellular stress. XC 2382 has a highly conserved GXGXXG signature sequence, that is a pyrophosphate binding motif found in NAD- and FAD- binding proteins, suggesting that it may bind pyrophosphate or nucleotide phosphates. ApaG also shares sequence homology with CorD, a Salmonella typhimurium protein associated with Co2+ sensitivity and Mg2+ homeostasis. The high degree of sequence conservation among ApaG homologs in bacteria indicated that it carries out some important biological functions. We have also assigned most of the resonances of the 1H, 15N, 13C nuclei using similar NMR experiments as described above. | en_US |