Construction of a Physiologically Active Photoaffinity Probe Based on the Structure of Bradykinin: Labelling of Angiotensin Converting Enzyme but Not Candidate Bradykinin Receptors on NG108–15 Cells

The peptides bradykinin and kallidin are released in response to noxious stimuli and mediate various physiological effects, including a direct stimulation of nociceptive afferent neurones. The nature of the receptor molecules through which these ligands act is presently unknown. We synthesised an iodinatable photoaffinity probe, N epsilon-4-azidosalicylylkallidin, and used it in an attempt to identify candidate bradykinin receptors on the NG108-15 neuroblastoma X glioma hybrid cell line. The ligand bound in subdued light to a particulate fraction of NG108-15 tumours and could be displaced by bradykinin with an IC50 of 0.33 nM. In a physiological assay, it behaved as an agonist equipotent with bradykinin. Gel analysis of the labelled products after photolysis of the iodinated ligand in the presence of NG108-15 cells or tumour membranes revealed bradykinin-blockable labelling of a glycoprotein with an Mr of 166,000. The probe was also able to label purified commercial angiotensin converting enzyme. The band labelled in NG108-15 cells was immunoprecipitable with a polyclonal antiserum to angiotensin converting enzyme, an enzyme shown to be present in low amounts in these preparations by direct binding using the iodinatable specific ligand MK351A.     

β-amyrin-induced apoptosis in Candida albicans triggered by calcium

The emergence of drug-resistant pathogens has urged researchers to discover alternatives for traditional antibiotics. β-amyrin, which is included in the category of triterpenoids extracted from plants, is known for its antimicrobial activity, although the underlying mechanism has not yet been revealed. This study was conducted to elucidate the antifungal mode of action of β-amyrin against Candida albicans. Based on the relevance between triterpenoids and oxidative molecules, reactive oxygen species (ROS) concentrations were detected, which showed a noticeable increment. Disruption of Ca²⁺ homeostasis in the cytosol was additionally analyzed, which was supported by interactions between two. Subsequently, decrease in mitochondrial membrane potential, increment of mitochondrial Ca²⁺, and ROS concentration were monitored, which suggested mitochondrial dysfunction modulated by Ca²⁺. Further investigation confirmed oxidative damage through glutathione reduction and DNA fragmentation. Accumulation of lethal damages resulted in the activation of caspases and externalization of phosphatidylserine, indicating the induction of yeast apoptosis by β-amyrin in C. albicans.     

Fungal ferritins: The ferritin from mycelia of Absidia spinosa is a bacterioferritin

Two distinct ferritin like iron containing proteins have been identified and isolated from the fungus Absidia spinosa; one from the spores and another from the mycelia. The mycelial protein has been purified and consists of two subunits of approx. 20 kDa. The N-terminal sequences of both subunits have been determined. The holoprotein as isolated contains approx. 750 iron atoms/molecule and exhibits a heme-like UV-Vis spectrum. Based on the heme spectrum and the high degree of sequence homology found, it has been established that the mycelial protein is a bacterioferritin. This is the first example demonstrating the presence of a bacterioferritin in a eukaryotic organism.