Cloning of a thermostable alpha-amylase gene from Thermomonospora curvata and its expression in Streptomyces lividans

The gene from Thermomonospora curvata CCM 3312 coding for thermostable alpha-amylase (tam) has been cloned in Streptomyces lividans TK 24 and localized to a 2.6 kb HindIII-BamHI fragment of DNA. The data presented here show that the tam gene is expressed at a high level in S. lividans and that the protein is efficiently excreted.     

Structure and function of the sterol carrier protein-2 N-terminal presequence

Although sterol carrier protein-2 (SCP-2) is encoded as a precursor protein (proSCP-2), little is known regarding the structure and function of the 20-amino acid N-terminal presequence. As shown herein, the presequence contains significant secondary structure and alters SCP-2: (i) secondary structure (CD), (ii) tertiary structure (aqueous exposure of Trp shown by UV absorbance, fluorescence, and fluorescence quenching), (iii) ligand binding site [Trp response to ligands, peptide cross-linked by photoactivatable free cholesterol (FCBP)], (iv) selectivity for interaction with anionic phospholipid-rich membranes, (v) interaction with a peroxisomal import protein [FRET studies of Pex5p(C) binding], the N-terminal presequence increased SCP-2's affinity for Pex5p(C) by 10-fold, and (vi) intracellular targeting in living and fixed cells (confocal microscopy). Nearly 5-fold more SCP-2 than proSCP-2 colocalized with plasma membrane lipid rafts and caveolae (AF488-CTB); 2.8-fold more SCP-2 than proSCP-2 colocalized with a mitochondrial marker (Mitotracker), but nearly 2-fold less SCP-2 than proSCP-2 colocalized with peroxisomes (AF488 antibody to PMP70). These data indicate the importance of the N-terminal presequence in regulating SCP-2 structure, cholesterol localization within the ligand binding site, membrane association, and, potentially, intracellular targeting.     

Positioning the nodule,the hormone dictum

The formation of a nitrogen-fixing nodule involves two diverse developmental processes in the legume root: infection thread initiation in epidermal cells and nodule primordia formation in the cortex. Several plant hormones have been reported to positively or negatively regulate nodulation. These hormones function at different stages in the nodulation process and may facilitate the coordinated development of the epidermal and cortical developmental programs that are necessary to allow bacterial infection into the developing nodule. In this paper, we review and discuss how the tissue specific nature of hormonal action dictates where, when and how a nodule is formed.Key words: nodulation, hormone regulation, epidermis, cortex     

Recognition of hypoxyloid and xylarioid <Emphasis Type="Italic">Entonaema</Emphasis> species and allied <Emphasis Type="Italic">Xylaria</Emphasis> species from a comparison of holomorphic morphology,HPLC profiles,and ribosomal DNA sequences

The genus Entonaema comprises Xylariaceae with hollow, gelatinous stromata that accumulate liquid. Some of its species, including the type species, appear related to Daldinia from a polyphasic approach, comprising morphological studies, comparisons of ribosomal DNA sequences, and high performance liquid chromatography (HPLC) profiles with diode array and mass spectrometric detection (HPLC-DAD-MS). This methodology was used to study Entonaema pallidum. Its major stromatal constituent was identified as xylaral, a secondary metabolite known from Xylaria polymorpha. This compound was detected in several Xylaria spp., including the tropical X. telfairii and morphologically similar taxa, whose stromata may also become hollow and filled with liquid. Cultures of E. pallidum resembled those of Xylaria, substantially differing from other Entonaema spp., in their morphology, 5.8S/ITS nrDNA sequences, and HPLC profiles. The type specimen of E. mesentericum was located in the spirit collection of the herbarium B and found to agree morphologically with the nomenclatorily younger E. pallidum. Traces of xylaral were even detected by HPLC-DAD-MS in the spirit in which the fungus had been preserved. Entonaema pallidum is thus regarded as a later synonym of E. mesentericum. Therefore, the latter name is transferred to Xylaria. A key to entonaemoid Xylariaceae is provided. Colour reactions (NH₃, KOH) of the ectostroma were applied to a limited number of Xylaria spp., but metabolite profiles of cultures appear more promising as chemotaxonomic traits to segregate this genus. As xylaral was also found in Nemania and Stilbohypoxylon spp., while being apparently absent in Hypoxylon and allied genera, it may be a chemotaxonomic marker for Xylariaceae with Geniculosporium-like anamorphs.     

Synthesis and cellular effects of cycloterpenals: cyclohexadienal-based activators of neurite outgrowth

An unusual class of diterpenoid natural products, 'cycloterpenals' (with a central cyclohexadienal core), that arise in nature by condensation of retinoids and other isoprenes, have been isolated from a variety of organisms including marine sponges as well as from the human eye. A milk whey protein has also demonstrated the formation of a cycloterpenal derived from beta-ionylidineacetaldehyde. Here, we generate a synthetic library of these molecules where we detail reaction conditions required to effect cross condensation of alpha,beta-unsaturated aldehydes as opposed to homodimerization. The ability of this class of molecules to activate neurite outgrowth activity is reported.