Optimization of the medium for the yeast synthesis of biotin

In order to increase the yield of biotin produced by the culture Sporobolomyces pararoseus, the medium containing sucrose, asparagine, MgSO4 (NH4)2SO4, KH2PO4, vitamin complex and trace elements was optimized. With the aid of a fractional factor experiment (2(5-1)) and a complete factor experiment (2(4)), the proportion of constituents was chosen in such a way as to double biotin yield, i.e. to increase it to 55.25 micrograms/l. An enrichment of the medium with yeast autolysate, casein hydrolysate and peptone in the presence of adenine increased biotin yield to 105.7 micrograms/l and cell productivity from 6.1 to 8.0 micrograms/l dry biomass.     

Micromechanical mapping of live cells by multiple-particle-tracking microrheology

This paper introduces the method of live-cell multiple-particle-tracking microrheology (MPTM), which quantifies the local mechanical properties of living cells by monitoring the Brownian motion of individual microinjected fluorescent particles. Particle tracking of carboxylated microspheres imbedded in the cytoplasm produce spatial distributions of cytoplasmic compliances and frequency-dependent viscoelastic moduli. Swiss 3T3 fibroblasts are found to behave like a stiff elastic material when subjected to high rates of deformations and like a soft liquid at low rates of deformations. By analyzing the relative contributions of the subcellular compliances to the mean compliance, we find that the cytoplasm is much more mechanically heterogeneous than reconstituted actin filament networks. Carboxylated microspheres embedded in cytoplasm through endocytosis and amine-modified polystyrene microspheres, which are microinjected or endocytosed, often show directed motion and strong nonspecific interactions with cytoplasmic proteins, which prevents computation of local moduli from the microsphere displacements. Using MPTM, we investigate the mechanical function of alpha-actinin in non-muscle cells: alpha-actinin-microinjected cells are stiffer and yet mechanically more heterogeneous than control cells, in agreement with models of reconstituted cross-linked actin filament networks. MPTM is a new type of functional microscopy that can test the local, rate-dependent mechanical and ultrastructural properties of living cells.     

DNMT1 and DNMT3B gene variants and their association with endometriosis in South Indian women

Background

Endometriosis is a multifactorial estrogen dependent gynecological disease characterized by implantation of functional endometrial tissue at ectopic positions. Though this disease is benign, it is associated with an increased risk of malignant transformation. Epigenetic disruptions like aberrant DNA methylation, resulting changes in gene expression capacity, are important in tumor progression and malignant cellular transformation. Therefore, variation in genes involved in DNA methylation might lead to disease susceptibility.

Purpose

To investigate the association between DNA methyl transferases (DNMT1 and DNMT3B) single nucleotide polymorphisms (SNPs) and the risk of endometriosis in South Indian women.

Methods

In the present study, we examined the genotypic and allele distribution of DNMT1 (rs10423341C/A, rs2228611G/Aandrs4804490C/A) and DNMT3B (rs1569686G/T) among the endometriosis patients (n?=?150) and controls (n?=?150). The genotypes were analyzed by polymerase chain reaction (PCR) and sequencing methods. Haplotype frequencies for multiple loci and the standardized disequilibrium coefficient (D?) for pairwise linkage disequilibrium (LD) were surveyed by Haploview Software.

Result

Significant increase in the frequencies of DNMT1 rs10423341 (P?=?0.04601), rs2228611 (P?=?0.00175) and DNMT3B rs1569686 (P?=?0.033) genotypes and alleles was observed in patients compared to controls. In addition, the frequency of A/A/C (P?=?0.0065) haplotype was significantly high in patients. But the DNMT1 (rs4804490) SNP did not show significant association with the disease.

Conclusion

The DNMT1 and DNMT3B polymorphism may constitute an inheritable risk factor for endometriosis in South Indian women. To the best of our knowledge there is no reported study on the association of polymorphisms in DNMT1 and DNMT3B with endometriosis risk.

     

Control of ammonium concentration in Escherichia coli fermentations

A control system has been devised for the maintenance of stable ammonium concentrations throughout a fedbatch fermentation. The control system is based on an ammonia gas-sensing electrode that monitors a pH-adjusted effluent stream from the fermentor. To overcome the time lag between the fermentor and the electrode, feedback control included metered flows of ammonium to both the fermentor and the electrode vessel. The system was used to study the growth of Escherichia coli B (ATCC 11303) at controlled ammonium concentrations of 5 to 200mM. Apparent specific growth rates, biomass and protein production, and glucose yields were essentially constant from 5 to 170mM. Above 170mM ammonium growth was inhibited. As ammonium concentration decreased from 170 to 5mM, ammonium yields increased from 1 to 24 g cell dry wt/g ammonium utilized. The results demonstrate that control of ammonium concentrations at levels so low that ammonium would be exhausted in batch fermentations can significantly increase overall ammonium yields.     

Molecular identification and properties of a light-insensitive rice catalase-B expressed in E. coli

Catalase plays a central role in plant stress responses but is highly susceptible to photoinhibition. A rice catalase-B protein avoiding photoinhibition was developed by mutagenesis of specific amino acids: Leu-189 to Trp-189 and His-225 to Thr-225 and then recombinantly expressed in E. coli. In addition, the site specific mutation also induced 2–2.5-fold increase in enzyme velocity with high affinity for its substrate and showed nearly a 3-fold lower K _m than the wild protein. These characteristic of mutated rice catalase-B is highly promising in transgenic research to increase plant productivity under stress conditions.     

Melatonin Does Not Modulate Testicular Responsiveness to Altered Photoperiods. A Study During Different Phases of the Annual Gonadal Cycle in Roseringed Parakeets (Psittacula krameri)

The roseringed parakeet has been shown to exhibit a variable testicular responsiveness to both altered photoperiodic regimens and to treatment with melatonin during different phases of the annual gonadal cycle. Adult male roseringed parakeets were held under either natural photoperiods (NP), or long photoperiods (LP; 16L 8D), or short photoperiods (SP; 8L 16D) for a total period of 90 days. From day 46 onward, half of the total birds in each group were administered with the vehicle of melatonin, and the other birds were injected daily in the afternoon with melatonin (25 µg/ 100 g body wt.) till the end of the experiment. An identical experimental schedule was followed during the four different (preparatory, progressive, pre-breeding, and breeding) phases of the annual testicular cycle. The testicular activities in various bird groups were evaluated by volumetric, gravimetric, histometric and karyometric measurements, and by quantitative histological studies. The findings revealed that exogenous melatonin may exert either a suppressive influence or none at all on the testicular functions in relation to the photoperiodic schedule as well as to the reproductive phase of the concerned bird, but in no case modulates gonadal responsiveness to artificially altered photoperiods.