Morgan, Hugh W.Klages, Karin Ursula2026-06-162026-06-161991https://hdl.handle.net/10289/18367The sulphur-dependent archaebacterial isolate ANI was studied in aspects of its ecology, physiology, and enzymology. Organisms closely resembling isolate ANI were enriched from New Zealand hot pools within a temperature range of 55-91°C and a pH range of 5.5- 9.0 and a sodium content above 0.5g/ℓ. The concentration of sodium ions was found to be a third parameter determining the presence or absence of these organisms. Isolate ANI is a neutrophilic, anaerobic extreme thermophile. In the laboratory, growth was observed within a pH range of pH 5.4 to 9.2 and a temperature range between 55 and 85°C. The organism is an obligate heterotroph with a requirement for peptone. In the presence of peptone, several amino acids, α-keto acids, glucose, lactose, casein, and starch stimulate growth. Sodium and sulphur are required for growth. Sodium can be replaced by lithium and sulphur by cystine and glutathione. Sodium is required for utilization of several amino acids and for synthesis of ATP. Presence of a Na⁺ gradient across the cytoplasmic membrane is essential for growth and ATP production. Ionophores affecting the Na⁺ gradient show a strong inhibitory effect, while a protonophore or a K⁺ ionophore exerted virtually no inhibition. In addition to a Na⁺ gradient the existence of either a H⁺ gradient or a membrane potential is required for growth and generation of ATP. The Na⁺ gradient is possibly generated via a Na⁺-translocating ATPase. Utilization of branched-chain amino acids in the presence of sulphur results in production of NH₄, CO₂, H₂, sulphide, and branched-chain fatty acids. When valine is utilized the major fatty acid end product is isobutyrate and with leucine or isoleucine it is isovalerate or 2-methylbutyrate (which could not be distinguished by analytical techniques used here). The metabolic pathway of branched-chain amino acids was investigated. The proposed common metabolic pathway for valine, leucine, and isoleucine involves four steps: deamination and production of the α-keto acid, oxidation of the α-keto acid and production of the acyl-CoA, production of the acyl phosphate from the acyl-CoA, and production of the fatty acid from the acyl phosphate, the latter step being coupled to production of ATP from ADP and inorganic phosphate. The presence of amino acid aminotransferase, α-keto acid oxidoreductase, and fatty acid kinase, which catalyze the first, second and fourth reaction respectively, was demonstrated in crude cell extracts, while the presence of phosphate acyltransferase, catalyzing the third step, was only suggested. Substrate-level phosphorylation is the major, if not the sole means of ATP generation in isolate ANI. Reducing equivalents generated by amino acid oxidation are transferred to a postulated hydrogenase and sulphur reductase respectively and sulphide and H₂ are produced. No inhibitory effect of H₂ could be demonstrated and reduction of sulphur as means for H₂ detoxification does not appear to occur in isolate ANI. Instead, H₂ and sulphide are produced concomitantly during growth. A type II restriction endonuclease of ANI, termed ‘ANi I’, was partially purified and its recognition sequence determined as CTAG. The cut site within the sequence is not known.enAll items in Research Commons are provided for private study and research purposes and are protected by copyright with all rights reserved unless otherwise indicated.Thesis