Biosynthesis of mycothiol: Elucidation of the sequence of steps in Mycobacterium smegmatis

Several members of the Actinomycetales, including the medically important mycobacteria, produce 1-D-myo-inosityl-2-(N-acetyl- L-cysteinyl)amino-2-deoxy-α-D-glucopyranoside (trivial name mycothiol) as their principal low-molecular-mass thiol. The pseudo-disaccharide component of mycothiol, 1-D-myo-inosityl- 2-amino-2-deoxy-α-D-glucopyranoside (α-D-GI), was synthesized by ligation of 1-D,L-2,3,4,5,6-penta-O-acetyl-myo-inositol to 3,4,6-tri-O-acetyl-2-deoxy-2-(2,4-dinitrophenylamino)-α-D-glucopyranysyl bromide to give, in the first instance, an isomeric mixture of α- and β-linked pseudo-disaccharides. The α-coupled D,D and D,L isomers, α-D-GI and α-L-GI respectively, were purified from the mixture by TLC, followed by removal of the protecting groups. A cell-free extract of Mycobacterium smegmatis catalysed the ligation of cysteine, acetate and α-D-GI in the presence of ATP and Mg2+ to form mycothiol, as judged by HPLC. When no acetate was added to the incubation mixture, an additional thiol accumulated. In the presence of [14C]acetate no radiolabel was recovered in this species, but only in mycothiol. The additional thiol was isolated as the bimane derivative, and 1H and 1H-1H COSY NMR spectra confirmed its identity as desacetylmycothiol. A more complete conversion of desacetylmycothiol into mycothiol was achieved in the presence of acetyl-S-CoA. These results indicate that the biosynthesis of mycothiol proceeds by the sequential addition of cysteine and acetate to α-D-GI. The inositol moiety appears to be an important determinant of specificity, since α-L-GI was poorly utilized.Articl

Cretaceous sea-surface temperature evolution: Constraints from TEX₈₆ and planktonic foraminiferal oxygen isotopes

It is well established that greenhouse conditions prevailed during the Cretaceous Period (~ 145–66 Ma). Determining the exact nature of the greenhouse-gas forcing, climatic warming and climate sensitivity remains, however, an active topic of research. Quantitative and qualitative geochemical and palaeontological proxies provide valuable observational constraints on Cretaceous climate. In particular, reconstructions of Cretaceous sea-surface temperatures (SSTs) have been revolutionised firstly by the recognition that clay-rich sequences can host exceptionally preserved planktonic foraminifera allowing for reliable oxygen-isotope analyses and, secondly by the development of the organic palaeothermometer TEX86, based on the distribution of marine archaeal membrane lipids. Here we provide a new compilation and synthesis of available planktonic foraminiferal δ18O (δ18Opl) and TEX86-SST proxy data for almost the entire Cretaceous Period. The compilation uses SSTs recalculated from published raw data, allowing examination of the sensitivity of each proxy to the calculation method (e.g., choice of calibration) and places all data on a common timescale. Overall, the compilation shows many similarities with trends present in individual records of Cretaceous climate change. For example, both SST proxies and benthic foraminiferal δ18O records indicate maximum warmth in the Cenomanian–Turonian interval. Our reconstruction of the evolution of latitudinal temperature gradients (low, ±48°, palaeolatitudes) reveals temporal changes. In the Valanginian–Aptian, the low-to-higher mid-latitudinal temperature gradient was weak (decreasing from ~ 10–17 °C in the Valanginian, to ~ 3–5 °C in the Aptian, based on TEX86-SSTs). In the Cenomanian–Santonian, reconstructed latitudinal temperature contrasts are also small relative to modern (< 14 °C, based on low-latitude TEX86 and δ18Opl SSTs minus higher latitude δ18Opl SSTs, compared with ~ 20 °C for the modern). In the mid-Campanian to end-Maastrichtian, latitudinal temperature gradients strengthened (~ 19–21 °C, based on low-latitude TEX86 and δ18Opl SSTs minus higher latitude δ18Opl SSTs), with cooling occurring at low-, middle- and higher palaeolatitude sites, implying global surface-ocean cooling and/or changes in ocean heat transport in the Late Cretaceous. These reconstructed long-term trends are resilient, regardless of the choice of proxy (TEX86 or δ18Opl) or calibration. This new Cretaceous SST synthesis provides an up-to-date target for modelling studies investigating the mechanics of extreme climates