Phosphorylation of Escherichia coli enolase

In vivo labeling of Escherichia coli JA200 pLC 11-8 resulted in 32P incorporation into enolase as demonstrated by immunoaffinity chromatography and electrophoresis followed by autoradiography. Complete acid hydrolysis, followed by thin layer chromatography was employed for determination of the phosphoamino acid residue. Comparison with phosphoamino acid standards resulted in the identification of a labeled residue corresponding to phosphoserine. In vitro labeling of cell extracts from glucose and acetate grown cells resulted in differential labeling of enolase. When specific radioactivities of in vivo labeled enolase were compared, 7 times more label was incorporated at late log phase in glucose grown cells than in late log acetate grown cells. At stationary phase, only 2.5 times more label was incorporated into glucose compared to acetate. When 32P-labeled enolase from glucose grown cells was subjected to treatment with potato acid phosphatase, dephosphorylation of the enzyme could be observed. Monitoring enzyme activity during the acid phosphatase treatment revealed a 70% decrease for the forward enzyme reaction, and a 3-fold increase, followed by a gradual decrease to almost zero, for the reverse enzyme reaction. Complete reversal of the changes in activity was possible by adding an aliquot of partially purified enolase kinase plus ATP.     

Diatoms in Acid Mine Drainage and Their Role in the Formation of Iron-Rich Stromatolites

Adverse conditions in the acid mine drainage (AMD) system at the Green Valley mine, Indiana, limit diatom diversity to one species, Nitzschia tubicola. It is present in three distinct microbial consortia: Euglena mutabilis-dominated biofilm, diatomdominated biofilm, and diatom-exclusive biofilm. E. mutabilis dominates the most extensive biofilm, with lesser numbers of N. tubicola, other eukaryotes, and bacteria. Diatom-dominated biofilm occurs as isolated patches containing N. tubicola with minor fungal hyphae, filamentous algae, E. mutabilis, and bacteria. Diatom-exclusive biofilm is rare, composed entirely of N. tubicola.

Diatom distribution is influenced by seasonal and intraseasonal changes in water temperature and chemistry. Diatoms are absent in winter due to cool water temperatures. In summer, isolated patchy communities are present due to warmer water temperatures. In 2001, the diatom community expanded its distribution following a major rainfall that temporarily diluted the effluent, creating hospitable conditions for diatom growth. After several weeks when effluent returned to preexisting conditions, the diatom biofilm retreated to isolated patches, and E. mutabilis biofilm flourished.

Iron-rich stromatolites underlie the biofilms and consist of distinct laminae, recording spatial and temporal oscillations in physicochemical conditions and microbial activity. The stromatolites are composed of thin, wavy laminae with partially decayed E. mutabilis biofilm, representing microbial activity and iron precipitation under normal AMD conditions. Alternating with the wavy layers are thicker, porous, spongelike laminae composed of iron precipitated on and incorporated into radiating colonies of diatoms. These layers indicate episodic changes in water chemistry, allowing diatoms to temporarily dominate the system.     

The effectiveness of traditional and sling exercise strength training in women

Strength training often combines closed-kinetic-chain exercises (CKCEs) and open kinetic-chain exercises (OKCEs). The CKCE may be more effective for improving performance in lower-body training. Recently, we reported upper-body CKCE (using a commercially available system of ropes and slings, Redcord AS, Staubo, Norway) was as effective as OKCE training for strength gains and that CKCE was more effective than OKCE for improving throwing performance. To our knowledge the effectiveness of a strength training program that uses exclusively CKCE is unknown. In this study, we examined the effectiveness of CKCE vs. OKCE strength training programs in women enrolled in an introductory strength training program. Twenty-six participants were randomized to OKCE (traditional exercises) or CKCE (sling-based exercises). Participants completed 6 sets per week for 13 weeks. Pre and posttraining evaluations included the following: 1 repetition maximum (1RM) leg and bench press; sling exercise push-ups; isokinetic dynamometry; lateral step-down test; and the Star Excursion Balance Test. Both groups significantly improved bench press (by an average of 4-6 kg) and leg press (by an average of 23-35 kg) (p < 0.001). There was a significant group × time interaction (p < 0.001) for sling exercise push-ups (OKCE pre = 5.5 ± 8.6, OKCE post = 6.1 ± 8.2, CKCE pre = 6.8 ± 6.0, CKCE post = 16.9 ± 6.6). Isokinetic measures of knee extension, knee flexion, shoulder internal rotation, and shoulder external rotation increased (improvements ranged from 2.7 to 27.7%), with no group differences. Both OKCE and CKCE strength training elicited similar changes in balance. We conclude that CKCE training is equally as effective as OKCE training during the initial phases of a strength training program in women. The fact that only CKCE improved sling exercise push-ups supports previous findings suggesting functional superiority of CKCE.     

低温对植物叶片中超氧物歧化酶、过氧化氢酶和过氧化氢水平的影响

番茄和鸡蛋果叶片中可提取的SOD活性不受低温的影响。在电泳谱带上SOD主同工酶带被氰化物而不被低温抑制,次同工酶带在低温下不稳定,且活性很低,它的变化不影响总的SOD活性。一些冷敏感植物叶片中CAT活性被低温抑制,而H_2O_3水平在低温下稳定或有增加,这可能使毒性更强的羟基离子(OH·)易于形成。     

Purification and characterization of enolase fromEscherichia coli

A method for the purification of enolase (EC 4.2.1.11) from an overproducing strain ofEscherichia coli JA 200 pLC 11–8 is described. The procedure included treatment of the crude sonic extract with protamine sulfate, followed by ammonium sulfate fractionation, hydrophobic interaction chromatography with phenyl Sepharose, HPLC ion exchange chromatography with a DuPont Sax column, and HPLC hydrophobic interaction chromatography with a Bio-Rad 5-PW column. The enzyme was purified to homogeneity as determined by silver staining of 10% sodium dodecylsulfate polyacrylamide gels. The native molecular weight ofE. coli enolase was found to be 92 kilodaltons and consisted of two subunits of identical molecular weight, 46 kilodaltons each. The isoelectric point was found to be 4.9.     

Binding Interaction between Tet(M) and the Ribosome: Requirements for Binding

Tet(M) protein interacts with the protein biosynthesis machinery to render this process resistant to tetracycline by a mechanism which involves release of the antibiotic from the ribosome in a reaction dependent on GTP hydrolysis. To clarify this resistance mechanism further, the interaction of Tet(M) with the ribosome has been examined by using a gel filtration assay with radioactively labelled Tet(M) protein. The presence of GTP and 5′-guanylyl imido diphosphate, but not GDP, promoted Tet(M)-ribosome complex formation. Furthermore, thiostrepton, which inhibits the activities of elongation factor G (EF-G) and EF-Tu by binding to the ribosome, blocks stable Tet(M)-ribosome complex formation. Direct competition experiments show that Tet(M) and EF-G bind to overlapping sites on the ribosome.     

Ribosomal RNA sequence phylogeny is not congruent with ascospore morphology among species in Ceratocystis sensu stricto

The genus Ceratocystis sensu stricto includes important fungal pathogens of woody and herbaceous plants. This genus is distinguished from species in Ceratocystis sensu lato by the presence of Chalara anamorphs. Ascospore shape has been used extensively in delineating Ceratocystis taxa, which show a large variety of ascospore shapes. Sequence analysis of one region of he 18S ribosomal RNA subunit and two regions of the 28S ribosomal RNA subunit showed that there was a majority of multiple substitutions at nucleotide sites and that there was a low transition/transversion ratio, T = 0.72. Both of these results suggest that these are well established, old species. Ascospore morphology, for the most part, was not congruent with the molecular phylogeny, and the use of morphological characters may be misleading in the taxonomy of these species.