Surface proteins of Gram-positive bacteria are covalently linked to the cell wall envelope by a mechanism requiring an N-terminal signal peptide and a C-terminal LPXTG motif sorting signal. We show here that surface proteins of Staphylococcus aureus arrive at two distinct destinations in the bacterial envelope, either distributed as a ring surrounding each cell or as discrete assembly sites. Pr...

Glutamine synthetase (GS) and glucose-6-phosphate isomerase (GPI) were identified as novel adhesive moonlighting proteins of Lactobacillus crispatus ST1. Both proteins were bound onto the bacterial surface at acidic pHs, whereas a suspension of the cells to pH 8 caused their release into the buffer, a pattern previously observed with surface-bound enolase and glyceraldehyde-3-phosphate dehydrog...

Glutamine synthetase (GS) type I is a key enzyme in nitrogen metabolism, and its activity is finely controlled by cellular carbon/nitrogen balance. In cyanobacteria, a reversible process that involves protein-protein interaction with two proteins, the inactivating factors IF7 and IF17, regulates GS. Previously, we showed that three arginine residues of IFs are critical for binding and inhibitio...

Glutamine synthetase (GS; EC 6.3.1.2) is the pivotal enzyme of nitrogen metabolism in prokaryotes. Control of bacterial GS activity by reversible adenylylation has provided one of the classical paradigms of signal transduction by cyclic cascades. By contrast, in the present work we show that cyanobacterial GS is controlled by a different mechanism that involves the interaction of two inhibitory...

Tandem affinity purification (TAP) has been widely used for the analysis of protein complexes. We investigated the parameters of the recently developed TAP method (GS-TAP) and its application in Drosophila. This new tag combination includes two Protein G modules and a streptavidin binding peptide (SBP), separated by one or two TEV protease cleavage sites. We made pMK33-based GS-TAP vectors to a...

The relative proportions and tissue distribution of the long (Gs[alpha]-L) and short (Gs[alpha]-S) variants of the a subunit of the stimulatory G-protein (Gs[alpha]) change under a wide range of metabolic conditions, such as cellular differentiation, ontogenetic development, ageing and various adaptive processes. Although the two variants of Gs(alpha) are generally regarded to be functionally i...

The cyanobacterial circadian oscillator can be reconstituted in vitro by mixing three clock proteins, KaiA, KaiB and KaiC, with ATP. KaiC is the only protein with circadian rhythmic activities. In the present study, we tracked the complex formation of the three Kai proteins over time using blue native (BN) polyacrylamide gel electrophoresis (PAGE), in which proteins are charged with the anionic...

To understand how glycogen affects skeletal muscle physiology, we examined enzymes essential for muscle glycogen synthesis and degradation using single fibers from quiescent and stimulated rat skeletal muscle. Presenting a shift in paradigm, we show these proteins are differentially associated with glycogen granules. Protein diffusibility and/or abundance of glycogenin, glycogen branching enzym...

Glycogen is a primary form of energy storage in eukaryotes that is essential for glucose homeostasis. The glycogen polymer is synthesized from glucose through the cooperative action of glycogen synthase (GS), glycogenin (GN), and glycogen branching enzyme and forms particles that range in size from 10 to 290 nm. GS is regulated by allosteric activation upon glucose-6-phosphate binding and inact...