Deletion and insertion of a 192-residue peptide in the active-site domain of glycosyl hydrolase family-2 beta-galactosidases

The monomeric multimetal-binding beta-galactosidase of Saccharopolyspora rectivirgula (srbg), a glycosyl hydrolase family-2 enzyme, has a unique sequence consisting of 192 amino acid residues with no similarity to known proteins. This 192-residue sequence (termed the "iota [iota] sequence") appears to be inserted into a sequence homologous to the active-site domain of the Escherichia coli lacZ enzyme (lacZbg). To assess the effects of the t sequence at specific sites of beta-galactosidase on the catalytic functioning and molecular properties of beta-galactosidase, deletion or insertion mutants of beta-galactosidases were constructed, expressed in LacZ- E. coli strains, and characterized: srbgdelta in which the iota sequence was deleted from srbg, and lacZbgI, in which the 192-residue iota sequence was inserted into the corresponding position (between Asp591 and Phe592) in the active-site domain of lacZbg. srbgdelta was a catalytically inactive, dimeric protein which retained multimetal-binding characteristics, suggesting that the iota sequence is very important for maintaining the structure necessary for the catalytic functioning and the monomeric structure of srbg but is not responsible for the unique metal ion requirements of srbg. On the other hand, lacZbgI existed as a mixture of a monomer, a tetramer, and higher multimers. The monomeric species was inactive, whereas the tetramer and other multimers were catalytically active (V(max )K(m) value, 25% of that of lacZbg) and highly specific for beta-D-galactoside. The tetrameric lacZbgI was activated by Mg2+ and Mn2+ with lowered metal affinities, and the stoichiometry of metal binding was unchanged from that of lacZbg. These results, along with the published stereo structure of lacZbg, suggest that, in lacZbgI, the inserted 192-residue iota peptide could fold independently of the lacZbg domains into a "sub-domain," lying distant from the active site and subunit interfaces.     

Novel sugar phosphotransferase system applicable to the efficient labeling of the compounds synthesized via the non-mevalonate pathway in Escherichia coli

It is known that 1-deoxy-D-xylulose taken up by Escherichia coli is used as the precursor of the compounds synthesized via the non-mevalonate pathway, such as isoprenoids, probably after conversion into 1-deoxy-D-xylulose 5-phosphate. In this report, we show that a novel phospho(enol) pyruvate-dependent phosphotransferase system catalyzes the uptake and phosphorylation of 1-deoxy-D-xylulose.     

Wind energy planning: Development and application of a site selection method for wind energy conversion systems (WECS)

The use of wind energy by means of its conversion to electricity involves a number of constrains such as economic, environmental, technical, legal, social and institutional requirements. Planning for wind energy should solve these issues in the planning process by encouraging opportunities and discouraging constraints associated with the use of wind energy. The opportunities and constraints significant at the regional level (i.e. approximately 10,000 km²) were identified, and a systematic method was developed to select sites for large WECS by incorporating the identified factors. These factors include: wind resource, proximity to load centres, proximity to tie-in points, and exclusive land-use areas. The developed method was applied to the western Massachusetts region, and the first results of the study have been acquired.     

HVEM study of crack-tip dislocations in Si crystals prepared by FIB and twin-blade cutting method

Crack-tip dislocations in silicon crystals have been examined by using high-voltage electron microscopy. Cracks were introduced by the Vickers indentation method at room temperature and the indented specimens were annealed at high temperatures to induce dislocations around crack tips under the presence of residual stress due to the indentation. A selected area around a crack tip was thinned by a focused ion beam (FIB) technique. Specimens were thinned in advance by a twin-blade cutting (TBC) method, which is a simple cutting process for saving FIB machine time. A combination of FIB and TBC can be a useful thinning procedure for the efficient preparation of transmission electron microscopy specimens. Characteristic dislocation structures were observed around the tip of a crack, aiding the elucidation of dislocation processes, which is essential to increase the fracture toughness of materials.     

Ultrashort X-ray pulse generation using subpicosecond electron linac

As a promising tool for ultrafast material analyses, we propose to utilize the X-ray pulse which may be generated in a quite simple manner using subpicosecond electron linacs. The properties of the X-ray were numerically studied with the EGS4 code. Verification of the X-ray generation was also conducted at the Nuclear Engineering Research Laboratory (NERL) linac and clear diffraction patterns of characteristic X-ray were obtained for typical single crystals.     

A High-Frequency Resonant Inverter Using Current-Vector Control Scheme and Its Performance Evaluations

The authors introduce a new configuration of resonant-type high-frequency inverter having inherent fast control response of the output power and variable-voltage variable-frequency (VVVF) capability. The circuit is composed of a parallel combination of two series-resonant inverters with common input and output terminals. Both inverter units are operated at synchronous frequency and with an adjustable phase difference from 0° to 180°, allowing control of the output power from full to null power, respectively. Operation of this inverter is explained and computer-simulated operating waveforms and characteristic curves are shown in terms of normalized control variables and circuit parameters. A prototype inverter using Power MOSFET modules has been originally tested with a high-frequency induction heating and melting load to demonstrate experimentally the proposed control principle and the steady-state inverter performances under parallel tuned load conditions.