An antibody microarray consists of antibody bound to a surface. Antibody microarrays have great potential in many fields, particularly as a tool to detect antigens. Unfortunately, antibodies suffer from poor performance. A greater understanding of how antibodies interact with surfaces would improve microarray design and performance, but experimental methods fall short of being able to observe these interactions. Therefore molecular simulation has emerged as the primary method to study protein/surface interactions.The simulations here were coarse grain simulations performed using the model of Karanicolas and Brooks. Additionally, an advanced surface model was used that allows for different surface chemistries. PyMBAR analysis was used to find heat capacities and determine relative stabilities of different linkers and tethering sites for the antibody/surface system.The actual work looked at how 24 different tethering sites affect antibody stability on two different surfaces and examined nine linkers varying in length and rigidity. Ultimately the findings were that antibody stability is a function of tethering position when tethered to a hydrophobic surface, but not when tethered to a hydrophilic surface. Furthermore, the length and rigidity of the linkers do not have a significant impact on stability.

Phosphite has been used to effectively control the soil borne plant pathogen Phytophthora cinnamomi in many horticultural crops, forest trees and natural ecosystems. However, the molecular mechanisms behind phosphite action on this pathogen are poorly understood. Several studies have shown that phosphite inhibits growth and zoospore production of P. cinnamomi and in addition induces significant physiological and metabolic changes in the mycelium. As an approach to understanding the mechanisms and relevance of these changes in the pathogen, the effect of phosphite on gene expression was investigated using microarray analysis. To construct the microarray, RNA was extracted from phosphite-treated (40 ug/ml) mycelium of P. cinnamomi isolate MP 80. The chosen phosphite concentration inhibited the mycelial growth by 70% but provided sufficient mycelium for RNA extractions after 4 days growth at 25C. The mRNA was reverse transcribed into cDNA and cloned into lambda to construct a library consisting of 2 million pfu of which 80 % were recombinant phage. The inserts were sequenced for a random selection of clones from the library. The nucleotide sequences generated revealed a range of different P. cinnamomi genes being expressed and demonstrated that the cDNA library provided a good representation of the transcripts expressed in P. cinnamomi. The types of genes found to be expressed in the mycelium of P. cinnamomi included genes encoding GTP binding proteins involved in vesicle transport, structural proteins involved in maintaining cell membrane integrity,elicitors, phosphatases and ribosomal proteins. Over nine thousand cDNA transcripts were randomly selected from the cDNA library and prepared by PCR amplification and purification for microarray construction. Custom made cDNA arrays containing 9216 cDNA transcripts were constructed and probed with RNA from untreated mycelium and mycelium grown in medium with 40 ug/ml phosphite. Two genes, EF-1 alpha and cinnamomin gene, identified by qRT-PCR as being constitutively expressed were also positioned on the arrays as positive controls. In the process of identifying constitutively expressed genes, qRT PCR revealed that phosphite down-regulated a gene encoding ubiquitin-conjugating enzyme, a component of the ubiquitin/proteasome pathway involved in the removal of abnormal and short lived-regulatory proteins and rate limiting enzymes. From the arrays a further seventy-two transcripts with altered patterns in gene expression (fold change > 2) were identified. The majority of the cDNA transcripts spotted on the array were down-regulated with changes in gene expression ranging from 2- to 3.5-fold. Thirty-two cDNA transcripts were up-regulated with changes in gene expression ranging from 2- to 16-fold. Characterisation by sequencing revealed that the most highly induced transcripts coded for ADP-ribosylation factors, an ABC cassette transporter and a glycosyl transferase. A transcript encoding a vitamin B6 biosynthesis protein was also identified as up-regulated by 2.9-fold. In contrast, the down-regulated transcripts coded for cellulose synthase I, annexin, glutamine synthetase, metallothionein and an alternative oxidase. The results are discussed in terms of possible roles and mechanism(s) of phosphite action within the mycelium of P.cinnamomi. This work is the first comprehensive screen for phosphite regulated-gene expression in P. cinnamomi and represents a significant step towards an understanding of the mode of action of phosphite on this organism. This thesis provides valuable information on the molecular interaction between phosphite and P. cinnamomi, which in future studies may stimulate the discovery of novel methods and cellular targets for the control of plant pathogenic Oomycetes.

Helicobacter pylori is a unique bacterial pathogen which colonises the human stomach. Infection with H. pylori has been linked to several disease outcomes including gastric and duodenal ulcer, gastric cancer and MALT lymphoma. Considering the harsh environment in which it resides and the lack of competition from other bacteria, this host/pathogen relationship is particularly interesting. Microarray analysis is a new and powerful technique which can be used to investigate various aspects of these complex interactions. Expression profiling of bacteria using microarrays remains in its infancy and thus appropriate methods were developed herein for investigating the transcriptional responses of H. pylori to various environments in vitro. Studies showed the tight relationship between growth phase dependent expression of iron homeostasis, motility and virulence genes in H. pylori for the first time. Consequently, the late exponential phase of growth was implicated as the most virulent growth phase of this bacterium in vitro. In response to mammalian cell co-culture, induced expression of H. pylori metabolism/respiration genes, genes of unknown function and genes encoding the 2-component regulators, HP1021 and HP0166, were detected. These represent a set of genes likely to be important specifically in the context of infection. To investigate the host response to infection a new mouse colonising strain of H. pylori, the Sydney Strain 2000 (SS2000), was isolated for use in comparative studies with the established strain, Sydney Strain 1 (SS1). Both host and strain specific effects were studied in a 15 month colonisation experiment using C57BL/6 and BALB/c mice. Genomic typing was used to investigate dynamic changes that occurred in the mouse-adapted strains during colonisation. In these animals reponses relating to the severity of inflammation and to the infecting H. pylori isolate were revealed by gene expression profiling. Previously unrealised cellular responses were uncovered. These included the significant down-regulation of both ferritin and haemoglobin expression. This perhaps suggests a mechanism for H. pylori induced iron deficiency anaemia. Physiological connections between colonisation, acid secretion and expression of the endocrine hormones were also implicated. These experiments have shown the utility of microarray analysis in the investigation of pathogenesis and have highlighted many directions for further investigation.

A methodology for the construction of predictive empirical models of physiological characteristics from microarray data is presented. The method, applied here to the study of the development of diabetes and insulin resistance, can be further expanded to other cases and to also include a variety of other data, such as protein expression, or metabolic flux data. The importance of several of the genes identified by the modeling methodology can be verified by comparison with results from prior literature. This implies potentially significant roles in diabetes for several of the uncharacterized genes discovered during the modeling procedure. / Singapore-MIT Alliance (SMA)

No description available.

Los glucosinolatos son compuestos cianogénicos naturales que se degradan por la acción de tioglucosidasas endógenas (mirosinasas) para dar lugar a isotiocianatos, tiocianatos y nitrilos; muchos de ellos han captado la atención por sus diversas propiedades biológicas, entre otras, como agentes preventores del cáncer, biopesticidas, antiafrodisiacos. Luego del secuenciamiento del genoma de Arabidopsis, se ha elucidado mejor la ruta de biosíntesis de los glucosinolatos, identificandose los primeros reguladores de la ruta, estudios de su función, así como relaciones evolutivas entre rutas parecidas. / Glucosinolates are natural cyanogenic compounds that are broken down by the action of endogenous tioglucosidases (mirosinases) to produce isothiocyanates, thiocyanate, and nitriles; many of them call the attention to researchers by diverse biological properties, such as cancer preventing, biopesticides, anti-aphrodisiacs. After the sequencing of Arabidopsis genome was completed, the glucosinolates biosynthetic process has been better elucidated, identifying the first regulators of the pathway, function research, and also the evolution relationship between similar pathways.

Thesis (M. Phil.)--Hong Kong University of Science and Technology, 2003. / Includes bibliographical references (leaves 103-111). Also available in electronic version. Access restricted to campus users.

Thesis (Ph.D.)--City University of Hong Kong, 2009. / "Submitted to Department of Biology and Chemistry in partial fulfillment of the requirements for the degree of Doctor of Philosophy." Includes bibliographical references (leaves 133-147)

Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 117-135).

Thesis (Ph.D.)--University of Nebraska-Lincoln, 2009. / Title from title screen (site viewed March 2, 2010). PDF text: 100 p. : col. ill. ; 953 K. UMI publication number: AAT 3379821. Includes bibliographical references. Also available in microfilm and microfiche formats.