Studies of the reduction of inorganic sulphate in the rumen of sheep

The ability of ruminants to utilize inorganic sulphur was first indicated by Warth (1932) and has been confirmed by many workers in recent years (Moir et al., 1967). The reduction of inorganic sulphate to sulphide has been demonstrated in the rumen (Lewis, 1954; Anderson, 1956) and in vitro (Henderickx, 1961) and inorganic sulphur amino acids of rumen microbial protein (Block et al., 1951; Henderickx 1961). Finally cysteine and methionine synthesized in the rumen have been isolated from tissue proteins of. the host animal (Block and Stekol, 1950; Kulwich et al., 1957). Although a vast quantity of literature exists concerning the biology of micro-organisms that oxidize and reduce inorganic sulphur compounds, until recently little was known about the enzymatic mechanisms involved in the metabolism of these compounds. A large and diverse group of micro-organisms reduces sulphate in the small amounts required for the synthesis of cellular material, as evidenced by the ability of these organisms to grow on sulphate as their sole source of sulphur. This small-scale reduction of sulphate has been termed assimilatory sulphate reduction (Postgate, 1959). A much smaller group of micro-organisms reduces sulphate in great excess of nutritional requirements and produces massive amounts of sulphide. This large scale reduction of sulphate to sulphide has been termed dissimilatory sulphate reduction (Postgate, 1959). The chemical reduction of sulphate is a relatively difficult reaction and little progress was made on the mechanisms of biological sulphate reduction until Robbins and Lipmann (1958) simultaneously with Wilson and Bandurski (1958) elucidated the mechanism of sulphate activation in a number of organisms. Their observations, along with the initial observations of De Melo and Wizerkanisk (1956) are summarized in reviews by Gregory and Robbins (1960) and Wilson (1962). However, little if any published work relates to the mechanisms of sulphate reduction in the rumen. This investigation was designed to show that the reduction of inorganic sulphate in the rumen of sheep followed a similar pathway as that of Robbins and Lipmann (1958) and Wilson and Bandurski (1958). The initial stages involved the isolation of the intermediates adenosine-5'-phosphosulphate (PAPS) and 3'-phospho -adenosine-5'- phosphosulphate (APS) from the rumen by use of S 35 -labelled sodium sulphate and the remainder of the experimental work was designed to show by use of the group V1 anions - molybdate, selenate, chromate and tungstate - that the enzyme ATP-sulphurylase was involved in the activation of sulphate by ATP prior to reduction. The review of literature covers the general sulphur metabolism and a further section discusses the bacterial sulphate reduction in more detail. The experimental section of this thesis is divided into two parts. In part A, the isolation of APS and PAPS is presented. In section B the preliminary responses of the group V1 anions on inorganic 4 sulphate reduction are presented followed by a more detailed study of the responses to these group V1 anions. Finally the behaviour of pure species Desulphovibris desulphuricans - and sulphate-reducing bacteria isolated from the rumen in response to group V1 anions has been investigated. Raw experimental data and statistical analyses have been recorded in the appendices