Evolution of an inducible penicillin-target protein in methicillin-resistant Staphylococcus aureus by gene fusion

A new beta-lactam-inducible penicillin-binding protein (PBP) that has extremely low affinity to penicillin and most other beta-lactam antibiotics has been widely found in highly beta-lactam(methicillin)-resistant Staphylococcus aureus (MRSA). The gene for this protein was sequenced and the nucleotide sequence in its promoter and close upstream area was found to show close similarity with that of staphylococcal penicillinase, while the amino acid sequence over a wide range of the molecule was found to be similar to those of two PBPs of Escherichia coli, the shape-determining protein (PBP 2) and septum-forming one (PBP 3). Probably the MRSA PBP (Mr 76462) evolved by recombination of two genes: an inducible type I penicillinase gene and a PBP gene of a bacterium, causing the formation of a beta-lactam-inducible MRSA PBP.     

New mutations fts-36, lts-33, and ftsW clustered in the mra region of the Escherichia coli chromosome induce thermosensitive cell growth and division.

Three new mutants of Escherichia coli showing thermosensitive cell growth and division were isolated, and the mutations were mapped to the mra region at 2 min on the E. coli chromosome map distal to leuA. Two mutations were mapped closely upstream of ftsI (also called pbpB), in a region of 600 bases; the fts-36 mutant showed thermosensitive growth and formed filamentous cells at 42 degrees C, whereas the lts-33 mutant lysed at 42 degrees C without forming filamentous cells. The mutation in the third new thermosensitive, filament-forming mutant, named ftsW, was mapped between murF and murG. By isolation of these three mutants, about 90% of the 17-kilobase region from fts-36-lts-33 to envA could be filled with genes for cell division and growth, and the genes could be aligned.     

Molecular cloning of the gene of a penicillin-binding protein supposed to cause high resistance to beta-lactam antibiotics in Staphylococcus aureus.

A novel penicillin-binding protein, PBP-2' (Mr about 75,000), is known to be induced in excessively large amount by most beta-lactam compounds in cells of a clinically isolated strain of Staphylococcus aureus, TK784, that is highly resistant to beta-lactams and also most other antibiotics. This protein has very low affinities to most beta-lactam compounds and has been supposed to be the cause of the resistance of the cells to beta-lactams. A 14-kilobase DNA fragment was isolated from the cells that carried the gene encoding this penicillin-binding protein and also a genetically linked marker that is responsible for the resistance to tobramycin. This DNA was cloned on plasmid pACYC184 and was shown to cause both production of PBP-2' and resistance to tobramycin in Escherichia coli cells. However, the formation of PBP-2' in E. coli was only moderate and was independent of normal inducer beta-lactams. The PBP-2' formed in the E. coli cells showed slow kinetics of binding to beta-lactams similar to that of PBP-2' formed in the original S. aureus cells and gave a similar pattern of peptides to the latter when digested with the proteolytic V8 enzyme of S. aureus.     

Sulfated extracellular polysaccharide production by the halophilic cyanobacterium Aphanocapsa halophytia immobilized on light-diffusing optical fibers

 The cyanobacterium, Aphanocapsa halo-phytia MN-11, was immobilized in calcium alginate gel and coated on light-diffusing optical fibers (LDOF) for sulfated extracellular polysaccharide production. Results indicated that sulfated extracellular polysaccharide production depends on the number of immobilized cells and the light intensity. In addition, the production rate reached 116.0 mg (mg dry cells)^-1 day^-1 when the cells that were immobilized on LDOF were incubated under a light intensity of 1380 cd sr m^-2 at a cell concentration of 1.0×10⁸ cells/cm³ gel. Cells immobilized on LDOF produced about ten times more sulfated extracellular polysaccharide than those immobilized in calcium alginate beads only (11.7 mg(mg dry cells)^-1day^-1). Received: 31 March 1995/Revised last revision 12 June 1995/Accepted 26 July 1995     

The 55-kilodalton protein in an oriC complex fraction is glycogen synthase.

Three proteins with molecular masses of 35, 55, and 75 kDa were found in an oriC complex fraction after purification through CsCl density gradient centrifugation (W. G. Hendrickson, T. Kusano, H. Yamaki, R. Balakrishnan, M. King, J. Murchie, and M. Schaechter, Cell 30:915-923, 1982). Of these three proteins, the 55-kDa protein was determined to be glycogen synthase on the basis of the N-terminal amino acid sequence and the molecular weight. The oriC complex was formed in glgA mutant cells, which produce no detectable glycogen, as well as in wild-type cells. None of the 35-, 55-, and 75-kDa proteins were detected in the fraction from this mutant. The results indicate that these proteins were not constituents of the oriC complex.     

ATPase activity of SopA, a protein essential for active partitioning of F plasmid

Summary The SopA, B, C genes of the F plasmid play an essential role in plasmid partitioning during cell division in Escherichia coli. In this paper, the products of the sopA and sopB genes were isolated and their biochemical activities studied. [-³²P]ATP was cross-linked to the SopA protein by UV irradiation; this cross-linking was observed only in the presence of magnesium ion, and was competitively inhibited in the presence of non-radioactive ATP, ADP and dATP, but not other NTPs or dNTPs. In contrast, no ATP binding activity was detected for the SopB protein. The SopA protein showed a modest magnesium ion-dependent ATPase activity and this activity was stimulated in the presence of DNA. The ATPase activity in the presence of DNA was further stimulated by addition of the SopB protein. However, the SopB protein alone failed to stimulate the ATPase activity.     

Cell surface changes accompanying aging in human diploid fibroblasts : III. Division age and senescence revealed by concanavalin A-mediated red blood cell adsorption

The relationship between cell size, [³H]thymidine incorporation capacity, and cell surface property of human diploid fibroblasts was investigated using the concanavalin A (ConA)-mediated red blood cell (RBC) adsorption assay. Small cells in late passage populations adsorbed RBCs well with the RBC coating method (in which ConA-coated RBCs are adsorbed to fibroblasts) as did large cells of this population, while small cells in early passage populations did not. The RBC adsorption capacity of rapidly dividing cells with this method differed among young, middle-aged and old cell populations. The results suggest that temporal cell size and [³H]thymidine incorporating capacity is not a measure of the division age of human diploid cells at the individual cell level. On the other hand, RBC adsorption with the fibroblast coating method (in which RBCs are adsorbed to ConA-coated fibroblasts) occurred to non-dividing cells of the populations. Thus, the increase in RBC adsorption with this method is considered to be a reflection of the increase in non-dividing cells at phase III. Our results support the hypothesis that RBC adsorption with the RBC and fibroblast-coating methods represents a cell surface marker for division age and senescence of human diploid cells, respectively, at the individual cell level.     

The Expression Pattern of Classical MHC Class I Molecules in the Development of Mouse Central Nervous System

Classical major histocompatibility complex (MHC) class I, first identified in the immune system, is also expressed in the developing and adult central nervous system (CNS). Although the MHC class I molecules have been found to be expressed in the CNS of different species, a necessary step to elucidate the temporal and spatial expression patterns of MHC class I molecules in the brain development has never been taken. Frozen sections were made from the brains of embryonic and postnatal C57BL/6 J mice, and the expression of H-2D^b mRNA was examined by in situ hybridization. Immunofluorescence was also performed to define the cell types that express H2-D^b in P15 mice. At E10.5, the earliest stage we examined, H2-D^b was expressed in neuroepithelium of the brain vesicles. From E12.5 to P0, H2-D^b expression was mainly located at cerebral cortex, neuroepithelium of the lateral ventricle, neuroepithelium of aquaeductus and developing cerebellum. From P4 to adult, H2-D^b mRNA was detected at olfactory bulb, hippocampus, cerebellum and some nerve nuclei. The major cell types expressing H-2D^b in P15 hippocampus, cerebral cortex and olfactory bulb were neuron. H2-K^b signal paralleled that of H2-D^b and the expression levels of the two molecules were comparable throughout the brain. The investigation of the expression pattern of H-2D^b at both embryonic and postnatal stages is important for further understanding the physiological and pathological roles of H2-D^b in the developing CNS.     

基于LUCC的生态系统服务空间化研究——以张掖市甘州区为例

生态系统服务(ES)评价为减缓或阻止人类活动导致的各种复杂生态问题提供了一种可能的方法,同时也有助于解决日益增加的各种土地利用冲突.以张掖市甘州区为例,运用一种新的空间化方法开展生态系统服务研究,并注重实现与土地利用研究的集成分析.首先根据研究区不同的群落生境和土地利用类型划分生态系统服务类型,然后检验不同土地利用类型和方式对各类生态系统服务供给的影响,最后分析该方法和数据的不确定性.结果表明:1)在灌区和甘州区两个尺度上,4类ES生产能力值均表现为:文化服务＞支持服务＞调节服务＞供给服务;2)2000-2009年,4类ES供给都呈递减趋势,同时,城镇用地、路网建设等人类活动驱动的土地利用方式迅速增加,整个绿洲农业区处于过度开发状态;3)该方法可操作性强,多学科的数据和知识分析是ES研究的难点.     

Forced expression of FLO11 confers pellicle-forming ability and furfural tolerance on Saccharomyces cerevisiae in ethanol production

We constructed a plasmid that expresses FLO11 encoding a cell surface glycoprotein of Saccharomyces cerevisiae under the control of a constitutive promoter. This plasmid conferred pellicle-forming ability on the non-pellicle-forming industrial strain of S. cerevisiae at the air–liquid interface of the glucose-containing liquid medium. The induced pellicle-forming cells exhibited tolerance to furfural, which is a key toxin in lignocellulosic hydrolysates, in ethanol production.