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Identification and Mapping of Self-Assembling Protein Domains Encoded by the Escherichia coli K-12 Genome by Use of Repressor Fusions. Leonardo Mariño-Ramírez, 2004.Self-assembling proteins and protein fragments encoded by the Escherichia coli genome were identified from E . coli K-12 strain MG1655 . Libraries of random DNA fragments cloned into a series of 
repressor fusion vectors were subjected to selection forimmunity to
infection by phage
.
Survivors were identified bysequencing the ends of the inserts, and
the fused protein sequencewas inferred from the known genomic
sequence . Four hundred sixty-threenonredundant open reading
frame-encoded interacting sequencetags [ISTs] were recovered from
sequencing 2,089 candidates.These ISTs, which range from 16 to 794
amino acids in length,were clustered into families of overlapping
fragments, identifyingpotential homotypic interactions encoded by
232 E . coli genes.Repressor fusions identified ISTs from
genes in every protein-basedfunctional category, but membrane
proteins were underrepresented.The IST-containing genes were
enriched for regulatory proteinsand for proteins that form
higher-order oligomers . Forty-eight[20.7%] homotypic proteins
identified by ISTs are predictedto contain coiled coils . Although
most of the IST-containinggenes are identifiably related to proteins
in other bacterialgenomes, more than half of the ISTs do not have
identifiablehomologs in the Protein Data Bank, suggesting that they
mayinclude many novel structures . The data are available online
at http://oligomers.tamu.edu/.
Genome-Wide Transcriptional Profiling Analysis of Adaptation of Bacillus subtilis to High Salinity. Leif Steil, 2003.The gram-positive soil bacterium Bacillus subtilis often faces increases in the salinity in its natural habitats . A transcriptional profiling approach was utilized to investigate both the initial reaction to a sudden increase in salinity elicited by the addition of 0.4 M NaCl and the cellular adaptation reactions to prolonged growth at high salinity (1.2 M NaCl) . Following salt shock, a sigB mutant displayed immediate and transient induction and repression of 75 and 51 genes, respectively . Continuous propagation of this strain in the presence of 1.2 M NaCl triggered the induction of 123 genes and led to the repression of 101 genes . In summary, our studies revealed (i) an immediate and transient induction of the SigW regulon following salt shock, (ii) a role of the DegS/DegU two-component system in sensing high salinity, (iii) a high-salinity-mediated iron limitation, and (iv) a repression of chemotaxis and motility genes by high salinity, causing severe impairment of the swarming capability of B . subtilis cells . Initial adaptation to salt shock and continuous growth at high salinity share only a limited set of induced and repressed genes . This finding strongly suggests that these two phases of adaptation require distinctively different physiological adaptation reactions by the B . subtilis cell . The large portion of genes with unassigned functions among the high-salinity-induced or -repressed genes demonstrates that major aspects of the cellular adaptation of B . subtilis to high salinity are unexplored so far . Improving the Thermostability of Raw-Starch-Digesting Amylase from a Cytophaga sp . by Site-Directed Mutagenesis. Rong-Jen Shiau, 2003.A heat-stable raw-starch-digesting amylase (RSDA) was generated through PCR-based site-directed mutagenesis . At 65°C, the half-life of this mutant RSDA, which, compared with the wild-type RSDA, lacks amino acids R178 and G179, was increased 20-fold . While the wild type was inactivated completely at pH 3.0, the mutant RSDA still retained 41% of its enzymatic activity . The enhancement of RSDA thermostability was demonstrated to be via a Ca2+-independent mechanism .
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