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Biomedical subjects

Richard M Napier

Publications and source records attributed to Richard M Napier.

8 recordsLinked to original sources
PubMed2006

Novel tertiary amine oxide surfaces that resist nonspecific protein adsorption.

Novel surfaces derivatized with tertiary amine oxides have been prepared and tested for their ability to resist nonspecific protein adsorption. The oxidation of tertiary amines supported on triazine units was carried out using mCPBA to give a format allowing conjugation of biologically active ligands alongside them. Adsorption to these surfaces was tested and compared to adsorption to a set of commercial and custom oligo-/poly(ethylene glycol) (OEG/PEG) supports by challenging them with a protein display library presented on bacteriophage lambda. The new class of amine oxide surfaces is found to compare favorably with the performance of the OEG/PEG supports in the prevention of nonspecific binding.

Adsorption↗
PubMed2006

Receptors for auxin: will it all end in TIRs?

The role of TIR1 in ubiquitination and regulated degradation of Aux/IAA transcription factors has been recognized for some years, but recent results have shown that TIR1 itself is also the binding site for auxin. The affinity and specificity of TIR1 match properties anticipated of a nuclear auxin receptor and we look at how they compare with the properties of ABP1. We also consider the mechanism of auxin action via TIR1 and the likelihood that the TIR1 family could account for all auxin responses. It seems likely that the TIR1 system can account for a large part of the repertoire of auxin-mediated responses, but maybe not all.

Arabidopsis↗
PubMed2005

TIRs of joy: new receptors for auxin.

Back-to-back papers have described the identification of a family of receptors for the plant hormone auxin. Most developmental processes in plants are dependent on auxin signalling making this discovery a landmark in the search for the mechanism of auxin action. The TIR1 gene translates into a protein with recognised motifs including an F-box domain and TIR1 forms part of an important ubiquitination complex that tags other proteins for degradation. Specific amongst the targets of TIR1 are a set of auxin-regulated transcription factors. The latest work has shown that TIR1 itself is also the binding site for auxin making it an auxin receptor with no requirement for a biochemical signalling cascade.

Binding Sites↗
PubMed2005

Auxins.

Auxin is a multifactorial phytohormone that is required for cell division. Fine gradients determine points of developmental change in time and space. It is associated intimately with the axiality of plant growth, and increasing doses lead to cell expansion or inhibition of cell expansion in different tissues. From embryonic patterning to fruit dehiscence every plant process has some involvement with auxin as a hormonal signal, including responses to wounding. Moreover, synthetic auxins have widespread uses as agrochemicals, particularly as selective herbicides. Despite the importance of auxin as a plant signal the pathways of its biosynthesis are still not clear. Much more is known about auxin perception and the mechanisms through which gene transcription is regulated. One receptor has been identified, and protein crystallography data has explained its auxin-binding capacity, but this is likely to control only a subset of auxin-mediated responses. Little is known of the signal transduction intermediates. A second receptor has been nominated and may be involved in controlling auxin-mediated gene transcription. A complex set of proteins comprising signalosome and proteasome contribute to the regulation of sets of transcription factors to confer regulation by derepression. A set of auxin transport proteins has been described with associated regulatory interactors, and these account for polar auxin flow and the control of auxin movements across cells, tissues, and around the plant. The gradients these transport systems build regulate the responses of growth and differentiation, including the plant's response to gravity. These areas are described and discussed by relating the physiology of the whole plant to the details of genetic and protein activities.

Indoleacetic Acids↗
PubMed2004

Two distinct signaling pathways participate in auxin-induced swelling of pea epidermal protoplasts.

Protoplast swelling was used to investigate auxin signaling in the growth-limiting stem epidermis. The protoplasts of epidermal cells were isolated from elongating internodes of pea (Pisum sativum). These protoplasts swelled in response to auxin, providing the clearest evidence that the epidermis can directly perceive auxin. The swelling response to the natural auxin IAA showed a biphasic dose response curve but that to the synthetic auxin 1-naphthalene acetic acid (NAA) showed a simple bell-shaped dose response curve. The responses to IAA and NAA were further analyzed using antibodies raised against ABP1 (auxin-binding protein 1), and their dependency on extracellular ions was investigated. Two signaling pathways were resolved for IAA, an ABP1-dependent pathway and an ABP1-independent pathway that is much more sensitive to IAA than the former. The response by the ABP1 pathway was eliminated by anti-ABP1 antibodies, had a higher sensitivity to NAA, and did not depend on extracellular Ca(2+). In contrast, the response by the non-ABP1 pathway was not affected by anti-ABP1 antibodies, had no sensitivity to NAA, and depended on extracellular Ca(2+). The swelling by either pathway required extracellular K(+) and Cl(-). The auxin-induced growth of pea internode segments showed similar response patterns, including the occurrence of two peaks in the dose response curve for IAA and the difference in Ca(2+) requirements. It is suggested that two signaling pathways participate in auxin-induced internode growth and that the non-ABP1 pathway is more likely to be involved in the control of growth by constitutive concentrations of endogenous auxin.

Calcium↗
PubMed2002

Crystal structure of auxin-binding protein 1 in complex with auxin.

The structure of auxin-binding protein 1 (ABP1) from maize has been determined at 1.9 A resolution, revealing its auxin-binding site. The structure confirms that ABP1 belongs to the ancient and functionally diverse germin/seed storage 7S protein superfamily. The binding pocket of ABP1 is predominantly hydrophobic with a metal ion deep inside the pocket coordinated by three histidines and a glutamate. Auxin binds within this pocket, with its carboxylate binding the zinc and its aromatic ring binding hydrophobic residues including Trp151. There is a single disulfide between Cys2 and Cys155. No conformational rearrangement of ABP1 was observed when auxin bound to the protein in the crystal, but examination of the structure reveals a possible mechanism of signal transduction.

Amino Acid Sequence↗
PubMed2002

Spermidine-binding proteins. Purification and expression analysis in maize.

Polyamine-binding proteins have been identified in a wide range of organisms, including mammals, yeasts, and bacteria. In this work, we have investigated specific spermidine binding to plant membrane proteins purified from microsomes of etiolated maize (Zea mays) coleoptiles. In the final purification step, specific spermidine-binding activity (K(d) 6.02 10(-7) M) was eluted from a HiTrapQ fast-protein liquid chromatography column at about 0.25 M NaCl, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the most active fraction showed a major polypeptide of about 60 kD and another copurifying 18-kD protein. Competition experiments, performed on HiTrapQ active fractions, confirmed the specificity of the binding. Upon Sephadex G-100 gel filtration, spermidine binding was associated almost exclusively with the 18-kD protein. On the basis of the N-terminal sequences, degenerate oligonucleotide probes were designed and used to isolate, by reverse transcriptase-polymerase chain reaction and polymerase chain reaction, cDNA fragments of about 1 kb for the 60-kD protein, and 0.9 kb for the 18-kD protein. Northern-blot analysis performed on etiolated coleoptiles and different tissues from 10-d-old maize plants indicated the presence of two different mRNAs of 1.7 and 0.7 kb. Southern-blot analysis indicated that the genes encoding the 60- and 18-kD proteins are probably derived from differential processing of the same precursor mRNA. Using rabbit polyclonal antibodies raised against these proteins, affinity purification and dot-blot experiments detected analogous membrane proteins in monocot and dicot plants.

Amino Acid Sequence↗