Comparison of mcl-Poly(3-hydroxyalkanoates) synthesis by different Pseudomonas putida strains from crude glycerol: citrate accumulates at high titer under PHA-producing conditions

Background

Eukaryotic ubiquitin and SUMO are frequently used as tags to enhance the fusion protein expression in microbial host. They increase the solubility and stability, and protect the peptides from proteolytic degradation due to their stable and highly conserved structures. Few of prokaryotic ubiquitin-like proteins was used as fusion tags except ThiS, which enhances the fusion expression, however, reduces the solubility and stability of the expressed peptides in E. coli. Hence, we investigated if MoaD, a conserved small sulfur carrier in prokaryotes with the similar structure of ubiquitin, could also be used as fusion tag in heterologous expression in E. coli.

Results

Fusion of MoaD to either end of EGFP enhanced the expression yield of EGFP with a similar efficacy of ThiS. However, the major parts of the fusion proteins were expressed in the aggregated form, which was associated with the retarded folding of EGFP, similar to ThiS fusions. Fusion of MoaD to insulin chain A or B did not boost their expression as efficiently as ThiS tag did, probably due to a less efficient aggregation of products. Interestingly, fusion of MoaD to the murine ribonuclease inhibitor enhanced protein expression by completely protecting the protein from intracellular degradation in contrast to ThiS fusion, which enhanced degradation of this unstable protein when expressed in E. coli.

Conclusions

Prokaryotic ubiquitin-like protein MoaD can act as a fusion tag to promote the fusion expression with varying mechanisms, which enriches the arsenal of fusion tags in the category of insoluble expression.     

Influence of Ecological and Social Factors on Body Mass of Wild Chimpanzees

The chimpanzees (Pan troglodytes schweinfurthii) of Gombe National Park, Tanzania, were weighed regularly over a period of 33 yr, resulting in 1286 measurements on 31 males and 26 females aged 2–43 yr. Female growth slowed at 10 yr and that of males at 13 yr. Median adult body mass is 39 kg for males and 31.3 kg for females. Body mass varied between years. Chimpanzees were heaviest during a period of frequent banana provisioning. They were also heavier when community range size was large and population density within the range was low. Chimpanzees were heavier in the wet than in the dry season and body mass tracked rainfall in the preceding mo except for May in which mass was anomalously low. Dominance rank is significantly correlated with body mass for females but not males. High-ranking individuals tended to maintain more stable mass. Variability in body mass was greater for young and old individuals than for prime adults.     

The N-acetyl-D-glucosamine repressor NagC of Vibrio fischeri facilitates colonization of Euprymna scolopes

To successfully colonize and persist within a host niche, bacteria must properly regulate their gene expression profiles. The marine bacterium Vibrio fischeri establishes a mutualistic symbiosis within the light organ of the Hawaiian squid, Euprymna scolopes. Here, we show that the repressor NagC of V. fischeri directly regulates several chitin- and N-acetyl-D-glucosamine-utilization genes that are co-regulated during productive symbiosis. We also demonstrate that repression by NagC is relieved in the presence of N-acetyl-D-glucosamine-6-phosphate, the intracellular form of N-acetyl-D-glucosamine. We find that gene repression by NagC is critical for efficient colonization of E. scolopes. Further, our study shows that NagC regulates genes that affect the normal dynamics of host colonization.     

Why do male chimpanzees defend a group range?

Male chimpanzees, Pan troglodytes, cooperate to defend a community range within which resident females range in smaller core areas. There has been debate over exactly what males are defending, whether mates, territory or both. One hypothesis holds that males are defending mates, and that an increase in community range size will lead directly to the acquisition of more females. However, males frequently attack females as well as males at the edge of the community range. We examined 18 years of observational data on the Gombe chimpanzees to determine the behaviour of males during extragroup encounters, and the consequences of changes in community range size on the number of adult females and indirect measures of food availability. Males were always aggressive to males from other communities, and often attacked adult females, especially those that were not sexually receptive, were older, and/or had more than one offspring. The number of females did not increase with range size, but several measures suggested an increase in food availability with range size. These measures include more time spent in large foraging parties, higher encounter rates with resident females, more encounters with sexually receptive females and higher female reproductive rates. These findings suggest that males defend a feeding territory for their resident females and protect them from sexual harassment. Although a large range may eventually attract more females, this is not an immediate consequence of range expansion. Male number was not correlated with community range size.     

Use of the weighted jackknife method to calculate the variance in cellular-specific protein secretion rate: application to monoclonal antibody secretion rate kinetics in response to osmotic stress

The specific secretion rate (q, mug protein secreted/viable cell-h) and its variance are very useful to compare the capability of cell lines for protein secretion. An assessment of specific secretion rate variability is also beneficial and important when the specific secretion rate is to be used as an on-line process parameter to monitor culture production behavior or for in-process decisionmaking. Experimental errors in mammalian cell culture (e.g., protein concentration measurement and cell counting) and estimation error in the method of calculating q contribute to the total variance of the specific secretion rate. Although the variance of q is essential for comparing the differences between cell lines and the response of the same cell line to different nutrient or environmental conditions, few methods for calculating the variance of the specific secretion rate have been reported. As a model system, we have used the weighted jackknife method and the delta method to calculate the variance in the specific secretion rate of a murine monoclonal antibody (q(mAb)) determined by a differential method. These methods were applied to calculate q(mAb) and its standard deviation to determine the change in q(mAb) kinetics during batch culture of the 9.2.27 hybridoma in response to growth in hyperosmotic media or osmotic stress. Without osmotic stress, during exponential growth in DMEM + 5% FBS spinner culture, the estimate of q(mAb) decreases at least threefold. Results indicate that the 9.2.27 hybridoma responds to hyperosmotic media (400 mOsm, 470 mOsm) by significantly reducing the degree of q(mAb) decrease in the exponential phase, thus maintaining a higher q(mAb) through the stationary phase. The trend of q(mAb) during the batch cultures studied is further confirmed by t-test. Osmotic stress is statistically shown to be able to alter significantly the hybridoma-specific mAb secretion kinetics during batch culture. Determination of the variance of specific secretion rate using the weighted jackknife method offers a powerful approach for establishing the confidence limits of specific protein secretion rate between cell cultures in different nutritional or osmotic environments. (c) 1996 John Wiley & Sons, Inc.     

Electron microscopic studies on the small intestinal mucosa of rats after mechanical intestinal obstruction and ischemia

Ultrastructural changes in the small intestine during the early phase following mechanical obstruction were compared with those after vascular ligation. In both experiments (the early phase of mechanical ileus and ischemia) intestinal epithelial cells at the tips of villi showed common features. One of the most significant changes was an alteration in microvilli, with fragmentation into vesicles, narrowing of apical microvilli and decrease in number. The other change was fatty degeneration of the epithelial cells, which was accompanied by vesiculation of smooth endoplasmic reticulum beneath the terminal webs, fat deposition and dilated Golgi complex containing fat. These observations suggest that in the early phase of mechanical ileus, ischemic damage plays an important role.