Antitumour activities of levans produced by Zymomonas mobilis strains

Levans produced by four Zymomonas mobilis strains showed antitumour activity against sarcoma 180 and Ehrlich carcinoma in Swiss albino mice. Levans from two strains (ZAP and CP4) had the highest effects. NMR analysis showed that the polymers were composed only of fructose units. The results suggested that the antineoplasic effect is associated to the polysaccharide molecular weight and that a particular molecular weight range may be responsible for this effect.     

Variation in the ribosomal internal transcribed spacers (ITS1 and ITS2) among eight taxa of the Mimulus guttatus species complex

The internal transcribed spacer region (ITS1 and ITS2) of the 18S-25S nuclear ribosomal DNA sequence and the intervening 5.8S region were sequenced from three individuals in each of eight taxa of the Mimulus guttatus species complex. Three discrete variants, or "types," of ITS sequences were found, among which 30%-40% of sites differed, compared with 1%-2% within types. Dot plots indicate that these types were not related by conspicuous rearrangements or inversions. More than one ITS type was often found in the same taxon, and two of three ITS types span species boundaries, indicating their presence prior to speciation. These ITS sequences showed essentially no positional homology with the nearest sequenced relative, tomato. In contrast, the 5.8S region was relatively unvaried, with 8 of 162 sites varied in the sample among all eight taxa. The phylogeny inferred by the most common ITS sequence type, rooted by the two other ITS types, agreed with isozymes in showing the distinctness of M. nudatus, M. laciniatus, and M. tilingii from the other five taxa.      

Variation in heat shock proteins within tropical and desert species of poeciliid fishes

The 70-kilodalton heat shock protein (hsp70) family of molecular chaperones, which contains both stress-inducible and normally abundant constitutive members, is highly conserved across distantly related taxa. Analysis of this protein family in individuals from an outbred population of tropical topminnows, Poeciliopsis gracilis, showed that while constitutive hsp70 family members showed no variation in protein isoforms, inducibly synthesized hsp70 was polymorphic. Several species of Poeciliopsis adapted to desert environments exhibited lower levels of inducible hsp70 polymorphism than the tropical species, but constitutive forms were identical to those in P. gracilis, as they were in the confamilial species Gambusia affinis. These differences suggest that inducible and constitutive members of this family are under different evolutionary constraints and may indicate differences in their function within the cell. Also, northern desert species of Poeciliopsis synthesize a subset of the inducible hsp70 isoforms seen in tropical species. This distribution supports the theory that ancestral tropical fish migrated northward and colonized desert streams; the subsequent decrease in variation of inducible hsp70 may have been due to genetic drift or a consequence of adaptation to the desert environment. Higher levels of variability were found when the 30- kilodalton heat shock protein (hsp30) family was analyzed within different strains of two desert species of Poeciliopsis and also in wild-caught individuals of Gambusia affinis. In both cases the distribution of hsp30 isoform diversity was similar to that seen previously with allozyme polymorphisms.      

柠檬酸合酶的分子生物学研究进展

柠檬酸合酶（citrate synthase,CS）是细胞内多种重要代谢途径的关键酶。CS可催化草酰乙酸和乙酰辅酶A之间的缩合反应生成柠檬酸和辅酶A。通常革兰氏阳性细菌、古菌以及真核细胞的CS为同源二聚体,而革兰氏阴性细菌的CS为同源六聚体。根据其在细胞内的定位不同,CS可分为线粒体CS、乙醛酸循环体CS、过氧化物酶体CS。这些同工酶在能量代谢、植物脂肪的代谢、脂肪酸的氧化及细胞解毒过程中起着重要作用。不同来源的CS空间结构、催化机制和动力学性质十分相似。针对其生化特性、空间结构特点、催化机制以及分子进化等研究进展进行综述。     

The effects of glutaredoxin and copper activation pathways on the disulfide and stability of Cu,Zn superoxide dismutase

Mutations in Cu,Zn superoxide dismutase (SOD1) can cause amyotrophic lateral sclerosis (ALS) through mechanisms proposed to involve SOD1 misfolding, but the intracellular factors that modulate folding and stability of SOD1 are largely unknown. By using yeast and mammalian expression systems, we demonstrate here that SOD1 stability is governed by post-translational modification factors that target the SOD1 disulfide. Oxidation of the human SOD1 disulfide in vivo was found to involve both the copper chaperone for SOD1 (CCS) and the CCS-independent pathway for copper activation. When both copper pathways were blocked, wild type SOD1 stably accumulated in yeast cells with a reduced disulfide, whereas ALS SOD1 mutants A4V, G93A, and G37R were degraded. We describe here an unprecedented role for the thiol oxidoreductase glutaredoxin in reducing the SOD1 disulfide and destabilizing ALS mutants. Specifically, the major cytosolic glutaredoxin of yeast was seen to reduce the intramolecular disulfide of ALS SOD1 mutant A4V SOD1 in vivo and in vitro. By comparison, glutaredoxin was less reactive toward the disulfide of wild type SOD1. The apo-form of A4V SOD1 was highly reactive with glutaredoxin but not SOD1 containing both copper and zinc. Glutaredoxin therefore preferentially targets the immature form of ALS mutant SOD1 lacking metal co-factors. Overall, these studies implicate a critical balance between cellular reductants such as glutaredoxin and copper activation pathways in controlling the disulfide and stability of SOD1 in vivo.     

Human glutaredoxin 3 forms [2Fe-2S]-bridged complexes with human BolA2

Human glutaredoxin 3 (Glrx3) is an essential [2Fe-2S]-binding protein with ill-defined roles in immune cell response, embryogenesis, cancer cell growth, and regulation of cardiac hypertrophy. Similar to other members of the CGFS monothiol glutaredoxin (Grx) family, human Glrx3 forms homodimers bridged by two [2Fe-2S] clusters that are ligated by the conserved CGFS motifs and glutathione (GSH). We recently demonstrated that the yeast homologues of human Glrx3 and the yeast BolA-like protein Fra2 form [2Fe-2S]-bridged heterodimers that play a key role in signaling intracellular iron availability. Herein, we provide biophysical and biochemical evidence that the two tandem Grx-like domains in human Glrx3 form similar [2Fe-2S]-bridged complexes with human BolA2. UV-visible absorption and circular dichroism, resonance Raman, and electron paramagnetic resonance spectroscopic analyses of recombinant [2Fe-2S] Glrx3 homodimers and [2Fe-2S] Glrx3-BolA2 complexes indicate that the Fe-S coordination environments in these complexes are virtually identical to those of the analogous complexes in yeast. Furthermore, we demonstrate that apo BolA2 binds to each Grx domain in the [2Fe-2S] Glrx3 homodimer forming a [2Fe-2S] BolA2-Glrx3 heterotrimer. Taken together, these results suggest that the unusual [2Fe-2S]-bridging Grx-BolA interaction is conserved in higher eukaryotes and may play a role in signaling cellular iron status in humans.     

Effects of long-term fixation on histological quality of undecalcified murine bones embedded in methylmethacrylate

While long-term fixation and storage of specimens is common and useful for many research projects, it is particularly important for space flight investigations where samples may not be returned to Earth for several months (International Space Station) or years (manned mission to Mars). We examined two critical challenges of space flight experimentation: the effect of long-term fixation on the quality of mouse bone preservation and the preservation of antigens and enzymes for both histochemical and immunohistochemical analyses, and how the animal/sample processing affects the preservation. We show that long-term fixation minimally affects standard histological staining, but that enzyme histochemistry and immunolabeling are greatly compromised. Further, we demonstrate that whole animal preservation is not as suitable as whole leg or stripped leg preservation for long-term fixation and all histological analyses. Overall, we recommend whole leg processing for long-term storage of bone specimens in fixative prior to embedding in plastic for histological examination.