Christopher H. Moore

Cysteine S-glycosylation, a new post-translational modification found in glycopeptide bacteriocins

O‐glycosylation is a ubiquitous eukaryotic post‐translational modification, whereas early reports of S‐linked glycopeptides have never been verified. Prokaryotes also glycosylate proteins, but there are no confirmed examples of sidechain glycosylation in ribosomal antimicrobial polypeptides collectively known as bacteriocins. Here we show that glycocin F, a bacteriocin secreted by Lactobacillus plantarum KW30, is modified by an N‐acetylglucosamine β‐O‐linked to Ser18, and an N‐acetylhexosamine S‐linked to C‐terminal Cys43. The O‐linked N‐acetylglucosamine is essential for bacteriostatic activity, and the C‐terminus is required for full potency (IC50 2 nM). Genomic context analysis identified diverse putative glycopeptide bacteriocins in Firmicutes. One of these, the reputed lantibiotic sublancin, was shown to contain a hexose S‐linked to Cys22.

Phosphorescence depolarization and the measurement of rotational motion of proteins in membranes.

Rotational relaxation times of the order of many microseconds are typical of the presumably uniaxial rotation of membrane proteins [ 11. So far measurements of rotational relaxation times have been limited to those proteins which are present at high occupancy either in the orginal membrane, as with the band-3 proteins of erythrocyte ghosts [2], or in highly purified fragments of membrane such as the acetylcholine receptor protein of the electric organ of Torpedo mamorata [3], or in reconstituted membranes [4]. This restriction arises from the relative insensitivity of the two measurement methods usually used, either saturation transfer EPR spectroscopy [5] or the decay of linear dichroism following flash photolysis of an attached triplet-forming probe such as eosin [6]. We describe here a method for determining rotational relaxation times

Action of a cell wall proteinase from Lactococcus lactis subsp. cremoris SK11 on bovine αs1-casein

1 ms) by measurement of the depolarization of laser flashinduced phosphorescence of erythrosin (tetraiodoflurorescein) attached to the protein of interest. This new method provides experimental realisation of many earlier suggestions [7,8]: it exceeds the sensitivity of the photodichroism method [6] by a factor of -lo’, and the saturation transfer method [S] by -104. We illustrate the use of our method by describing the slow isotropic rotation of proteins in viscous media, and the anisotropic rotation of Ca”dependent ATPase in sarcoplasmic reticulum membrane.

[13] Phosphofructokinase from Streptococcus lactis

SummaryThe cell wall-associated proteinase from Lactococcus lactis subsp. cremoris SK11 was partially purified and incubated with αs1-casein for various times up to 48 h. Sixteen trifluoroacetic acid-soluble oligopeptide hydrolysis products were identified by determination of the aminp acid sequence. Eleven of these oligopeptides originated from the 78-residue sequence comprising the C-terminal region of αs1-casein and were present among the products after the first 60 min of digestion. Three oligopeptides from the N-terminal region and two others from the central region of the αs1-casein sequence were also present among the early digestion products although in smaller amounts than most of the oligopeptides from the C-terminal region. No cleat consensus sequence of amino acid residues surrounding the cleavage sites could be identified.

Incorporating Ethical and Trust-based Behaviour into Economics: A Transaction Cost Exploration

Publisher Summary This chapter describes an assay method of phosphofructokinase, purification of phosphofructokinase from Streptococcus lactis , and properties of this enzyme. Phosphofructokinase activity can be coupled via the enzymes fructose 1,6-bisphosphate aldolase, triosephosphate isomerase, and glycerol 3-phosphate dehydrogenase to NADH oxidation. The enzyme activity can then be determined spectrophotometrically by measuring the decrease in absorbance at 340 nm. The purification steps involved are growth of bacteria, preparation of crude extract, ammonium sulfate fractionation, chromatography on Blue Dextran–Sepharose, and crystallization. Streptococcus lactis phosphofructokinase appears to be a tetramer of identical subunits and the native enzyme has a molecular weight (MW) of 144,500. The purified enzyme can be stored at -12 ° C in Tris-glycerol buffer for up to six months with only 10–20% loss in activity. The enzyme has a pH optimum of 7.2–7.8 in a number of different buffers, but in phosphate buffer, both the pH optimum and the maximum velocity are lowered. The chapter indicates that use of the Blue Dextran–Sepharose affinity column is a very effective purification step in the isolation of S. lactis phosphofructokinase giving high yields of homogeneous enzyme.

An approach to the semisynthesis of acyl carrier protein

This paper explores ethical activity within a transaction cost (TC) framework. It discusses the theoretical and practical links between TC economisation and ethical and trust-based behaviour. By engendering a climate of trust at the micro/firm level, ethical behaviour can reduce TCs associated with monitoring and protection against broken contracts. At the macro/societal level, considerations such as stakeholders’ increasing ethical expectations, may also have TC implications. To distinguish these latter TCs from those at the more micro level, the paper terms them social transaction costs (STCs). While it might be possible to internalise TCs to achieve economisation, STCs cannot be reduced through internalisation.

Identification of alcaligin as the siderophore produced by Bordetella pertussis and B. bronchiseptica.

Abstract A scheme has been devised for the preparation of semisynthetic derivatives of acyl carrier protein (ACP). Acetylated synthetic ACP1–6 is coupled via its activated pentachlorophenol ester to native ACP (7–77), which had previously been acetylated and converted to the S-5′-dithiobis(2-nitrobenzoate)(DTNB) derivative. Removal of the DTNB moiety after the coupling yielded active ACP in good yield.