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71.
K. N. Faber S. Westra H. R. Waterham I. Keizer-Gunnink W. Harder G. AB M. Veenhuis 《Applied microbiology and biotechnology》1996,45(1-2):72-79
We describe the synthesis and purification of two functional peptides, namely human insulin-like growth factor II (IGF-II)
and Xenopus laevis magainin II in Hansenula polymorpha after their synthesis as hybrid proteins fused to the C terminus of endogenous amine oxidase. The hybrid genes, placed under
control of the H. polymorpha alcohol oxidase promoter (PAOX), were integrated into the genomic alcohol oxidase locus, yielding stable production strains. High-level synthesis of the
fusion proteins, exceeding 20% of total cellular protein, was obtained when the transformed strains were grown in methanol-limited
chemostat cultures; when expressed by itself, i.e. in the absence of the amine oxidase gene, IGF-II could not be recovered
from crude cell extracts, probably as a result of rapid proteolytic degradation. Accumulation in peroxisomes did not significantly
affect the IGF-II protein stability when expressed in the absence of the carrier protein. Apparently, fusion to the large
(±78 kDa) amine oxidase carrier particularly stabilizes the peptides and prevents them from proteolysis. After partial purification,
the fusion partners were readily separated by factor Xa treatment.
Received: 16 June 1995 / Accepted: 20 September 1995 相似文献
72.
S Jahan S Singh A Srivastava V Kumar D Kumar A Pandey CS Rajpurohit AR Purohit VK Khanna AB Pant 《Molecular neurobiology》2018,55(4):2828-2839
73.
We describe a new scaffold-free three-dimensional (3D) cell culture model using cholesteryl ester based lyotropic liquid crystal (LC) substrates. Keratinocytes were deposited randomly on the LC surface where they self-assembled into 3D microtissues or keratinospheroids. The cell density required to form spheroids was optimized. We investigated cell viability using dead/live cell assays. The adhesion characteristics of cells within the microtissues were determined using histological sectioning and immunofluorescence staining. Fourier transform infrared spectroscopy (FTIR) was used to characterize the biochemistry of the keratinospheroids. We found that both cells and microtissues could migrate on the LC surface. The viability study indicated approximately 80% viability of cells in the microtissues up to 20 days of culture. Strong intercellular adhesion was observed in the stratification of the multi-layered microspheroids using field emission-scanning electron microscopy (FE-SEM) and histochemical staining. The cytoskeleton and vinculins of the cells in the microtissues were expressed diffusely, but the microtissues were enriched with lipids and nucleic acids, which indicates close resemblance to the conditions in vivo. The basic 3D culture model based on LC may be used for cell and microtissue migration studies in response to cytochemical treatment. 相似文献
74.
Hannah R. Dykstra Stephanie R. Weldon Adam J. Martinez Jennifer A. White Keith R. Hopper George E. Heimpel Mark K. Asplen Kerry M. Oliver 《Applied and environmental microbiology》2014,80(18):5818-5827
Many insects are associated with heritable symbionts that mediate ecological interactions, including host protection against natural enemies. The cowpea aphid, Aphis craccivora, is a polyphagous pest that harbors Hamiltonella defensa, which defends against parasitic wasps. Despite this protective benefit, this symbiont occurs only at intermediate frequencies in field populations. To identify factors constraining H. defensa invasion in Ap. craccivora, we estimated symbiont transmission rates, performed fitness assays, and measured infection dynamics in population cages to evaluate effects of infection. Similar to results with the pea aphid, Acyrthosiphon pisum, we found no consistent costs to infection using component fitness assays, but we did identify clear costs to infection in population cages when no enemies were present. Maternal transmission rates of H. defensa in Ap. craccivora were high (ca. 99%) but not perfect. Transmission failures and infection costs likely limit the spread of protective H. defensa in Ap. craccivora. We also characterized several parameters of H. defensa infection potentially relevant to the protective phenotype. We confirmed the presence of H. defensa in aphid hemolymph, where it potentially interacts with endoparasites, and performed real-time quantitative PCR (qPCR) to estimate symbiont and phage abundance during aphid development. We also examined strain variation of H. defensa and its bacteriophage at multiple loci, and despite our lines being collected in different regions of North America, they were infected with a nearly identical strains of H. defensa and APSE4 phage. The limited strain diversity observed for these defensive elements may result in relatively static protection profile for this defensive symbiosis. 相似文献
75.
Siti Mutripah Maria Dyah Nur Meinita Ji-Young Kang Gwi-Taek Jeong AB Susanto Romanus Edy Prabowo Yong-Ki Hong 《Journal of applied phycology》2014,26(1):687-693
Seaweeds, particularly species of red macroalgae, are promising resources for bioethanol production because of their exceptionally high carbohydrate content. Of 20 seaweeds evaluated, Palmaria palmata (Rhodymenia palmata) contained the highest carbohydrate content (469.8 mg g?1 seaweed) with a carrageenan content of 354 mg g?1 seaweed. Such a high carrageenan content makes the high-volume production of bioethanol feasible. Acid hydrolysis of P. palmata in 0.4 M H2SO4 at 125 °C for 25 min released 27 mg of glucose, 218.4 mg of reducing sugars, and 127.6 mg of galactose per gram of seaweed. Ethanol fermentation of these hydrolysis products using an inoculum concentration of 1.5 mg mL?1 at 30 °C and 72 h in a shaking incubator at 130 rpm yielded 17.3 mg of ethanol per gram of seaweed. 相似文献
76.
运用16种酶蛋白编码的23个遗传座位对突尼斯非洲跳鼠(Jaculus jaculus)和埃及跳鼠(J.orientalis)自然群体的遗传变异和分化进行了电泳分析.结果表明,与其他啮齿动物等哺乳动物的相关数据比较,发现这两个种群体的遗传变异水平较低.非洲跳鼠群体的观测杂合度(Hobs)为0.08-0.19,多态座位百分比(P)为26.2%-45.2%,每个座何的平均等位基因数(A)为1.1-1.4;埃及跳鼠的Hobs为0.10-0.15,P为29.3%-44.1%,A为1.1-1.7.两个种群体各自的遗传分化程度较低(非洲跳鼠和埃及跳鼠的Fst分别为0.0017和0.0019).而两个种群体间的Fst为0.607(P<0.05),表明两个种之间高度的遗传分化.本研究支持这两个种分类地位的合法性,并强调了地理因素(环境类犁和生物气候阶段)对两个种遗传结构的影响. 相似文献
77.
M?nica G Campiteli Frederico M Soriani Iran Malavazi Osame Kinouchi Carlos AB Pereira Gustavo H Goldman 《BMC bioinformatics》2009,10(1):270
Background
Microarray techniques have become an important tool to the investigation of genetic relationships and the assignment of different phenotypes. Since microarrays are still very expensive, most of the experiments are performed with small samples. This paper introduces a method to quantify dependency between data series composed of few sample points. The method is used to construct gene co-expression subnetworks of highly significant edges. 相似文献78.
Patrícia Picciarelli-Lima André G Oliveira Adelina M Reis Evanguedes Kalapothakis Germán AB Mahecha Rex A Hess Cleida A Oliveira 《Reproductive biology and endocrinology : RB&E》2006,4(1):51-10
Background
Fluid homeostasis is critical for normal function of the male reproductive tract and aquaporins (AQP) play an important role in maintenance of this water and ion balance. Several AQPs have been identified in the male, but their regulation is not fully comprehended. Hormonal regulation of AQPs appears to be dependent on the steroid in the reproductive tract region. AQP9 displays unique hormonal regulation in the efferent ductules and epididymis, as it is regulated by both estrogen and dihydrotestosterone (DHT) in the efferent ductules, but only by DHT in the initial segment epididymis. Recent data have shown that a metabolite of DHT, 5-alpha-androstane-3-beta-17-beta-diol (3-beta-diol), once considered inactive, is also present in high concentrations in the male and indeed has biological activity. 3-beta-diol does not bind to the androgen receptor, but rather to estrogen receptors ER-alpha and ER-beta, with higher affinity for ER-beta. The existence of this estrogenic DHT metabolite has raised the possibility that estradiol may not be the only estrogen to play a major role in the male reproductive system. Considering that both ER-alpha and ER-beta are highly expressed in efferent ductules, we hypothesized that the DHT regulation of AQP9 could be due to the 3-beta-diol metabolite. 相似文献79.
The endoplasmic reticulum (ER) of specialized cells can undergo dramatic changes in structural organization, including formation of concentric whorls. We previously reported that depletion of Yip1A, an integral membrane protein conserved between yeast and mammals, caused ER whorl formation reminiscent of that seen in specialized cells. Yip1A and its yeast homologue Yip1p cycle between the ER and early Golgi, have been implicated in a number of distinct trafficking steps, and interact with a conserved set of binding partners including Yif1p/Yif1A and the Ypt1/Ypt31 Rab GTPases. Here, we carried out a mutational analysis of Yip1A to obtain insight into how it regulates ER whorl formation. Most of the Yip1A cytoplasmic domain was dispensable, whereas the transmembrane (TM) domain, especially residues within predicted TM helices 3 and 4, were sensitive to mutagenesis. Comprehensive analysis revealed two discrete functionally required determinants. One was E95 and flanking residues L92 and L96 within the cytoplasmic domain; the other was K146 and nearby residue V152 within the TM domain. Notably, the identified determinants correspond closely to two sites previously found to be essential for yeast viability (E76 and K130 in Yip1p corresponding to E95 and K146 in Yip1A, respectively). In contrast, a third site (E89) also essential for yeast viability (E70 in Yip1p) was dispensable for regulation of whorl formation. Earlier work showed that E76 (E95) was dispensable for binding Yif1p or Ypt1p/Ypt31p, whereas E70 (E89) was required. Collectively, these findings suggest that the ability of Yip1A to bind its established binding partners may be uncoupled from its ability to control ER whorl formation. In support, Yif1A knockdown did not cause ER whorl formation. Thus Yip1A may use the sites identified herein to interact with a novel binding partner to regulate ER membrane organization. 相似文献
80.
Lonneke Smeding Jan Willem Kuiper Frans B Pl?tz Martin CJ Kneyber AB Johan Groeneveld 《Respiratory research》2013,14(1):92