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IntroductionTo examine the relationship between the plasma glucose concentration (PG) and the pathways of hepatic glucose production (HGP), five groups of conscious rats were studied after a 6-h fast: (a) control rats (PG = 8.0±0.2 mM); (b) control rats (PG = 7.9±0.2 mM) with somatostatin and insulin replaced at the basal level; (c) Overproduction of glucose is the major cause of fasting hyperglycemia in both insulin-dependent (1) and non-insulin-dependent diabetes mellitus (NIDDM) (2).' Gluconeogenesis composes an higher percentage of hepatic glucose production (HGP) in diabetic as compared with nondiabetic individuals (3-6), and it has been postulated to be the "primary" determinant ofincreased HGP in diabetes (5). However, recent experimental evidence suggests that changes in the rates offormation of hepatic glucose-6-phosphate (G6P) through gluconeogenesis do not alter HGP in conscious dogs (7), healthy volunteers (8-10), and NIDDM subjects ( 1 1). The "final common pathway" for the net release of glucose into the circulation is regulated by the balance offluxes through glucokinase and glucose-6-phosphatase (G6Pase). The assessment of in vivo metabolic fluxes contributing to G6P formation and to hepatic glucose output, in combination with the steady-state changes in the hepatic G6P concentration, may allow one to discern ifthe key regulatory site for HGP is the formation of G6P or its net dephosphorylation to glucose. See Fig. 1 for a schematic representation. Soskin and Levine ( 12) first proposed that mammalian liver can rapidly change its glucose output in response to changes in the circulating glucose concentration independently from hormonal signals. It is now recognized that the plasma glucose concentration per se regulates HGP ( 13-18) and as much as 50% of the decline in plasma glucose concentration after glucose administration may be due to the combined effect of hyperglycemia per se on glucose disposal and HGP (18). However, relatively little information is available about the mechanism(s) by which hyperglycemia, independent of insulin, inhibits HGP. Basal HGP is markedly increased in some diabetic states despite the presence of hyperglycemia and normo-or hyperinsulinemia, both ofwhich are known to suppress HGP (2, 13-18). This may suggest a role for defective glucose-induced suppression ofHGP in the pathophysiology of fasting hyperglycemia in diabetes.HGP is composed of glycogen mobilization and gluconeogenesis. However, under postabsorptive conditions, i.e., basal insulin, a sizeable portion of the flux through G6Pase also comes from glucose cycling ( 19-23). Although hyperglycemia per se inhibits HGP, such an effect has not been demonstrated for total glucose output (TGO) (21 ), i.e., flux through G6Pase. Thus, it is possible that a component ofthe effect ofhyperglycemia per se in suppressing HGP is mediated by the enhanced 1. Abbreviations used in thispaper: G6P, glucose-6-phosphate; G6Pase, glucose-6-phosphatase; GC, glucose cycling; HGP, hepatic glucose production; NIDDM, non-insu...

The M2 protein from influenza A is a pH-activated proton channel that plays an essential role in the viral life cycle and serves as a drug target. Using spin labeling EPR spectroscopy we studied a 38-residue M2 peptide spanning the transmembrane region and its C-terminal extension. We obtained residue-specific environmental parameters under both high and low pH conditions for nine consecutive C-terminal sites. The region forms a membrane surface helix at both high and low pH although the arrangement of the monomers within the tetramer changes with pH. Both electrophysiology and EPR data point to a critical role for residue Lys 49.M2 is a 96-residue homotetrameric integral membrane protein with a small N-terminal ectodomain, a single transmembrane helix and a C-terminal cytoplasmic tail. Despite data from solid state NMR (1), x-ray crystallography (2) and solution NMR (3), a detailed understanding of how the M2 protein works continues to puzzle investigators and generate sharp controversy.The majority of published studies on the proton channel function of M2 have focused on the transmembrane (TM) 1 domain. However, truncation studies indicate that the cytoplasmic domain also plays a role in channel stability (4). Proteolysis of micelle-bound full length M2 revealed that a 15-20 residue segment C-terminal to the TM helix was highly protected from cleavage by proteases (5). Helical wheel analysis of the protected region (5) suggested that the segment could form an amphiphilic helix, consistent with later findings from solid state NMR on M2 protein in lipid bilayers (6). In order to further test the proposed models, we probed the conformation of the segment C-terminal to the TM domain at both high and low pH using sitedirected spin-labeling (SDSL) and electron paramagnetic resonance (EPR) spectroscopy.EPR studies were performed on a series of 38-residue synthetic M2 peptides (residues 23-60; M2TMC) spanning the TM region and the beginning of the C-terminal domain. We spin- † This research was supported by R01AI57363 (LHP), GM56423 (WFD), a Henry Dreyfus Teacher Scholar Award (KPH) and R15AI074033 (KPH).

Maintenance of periodontal health or transition to a periodontal lesion reflects the continuous and ongoing battle between the vast microbial ecology in the oral cavity and the array of resident and emigrating inflammatory/immune cells in the periodontium. This war clearly signifies many 'battlefronts' representing the interface of the mucosal-surface cells with the dynamic biofilms composed of commensal and potential pathogenic species, as well as more recent knowledge demonstrating active invasion of cells and tissues of the periodontium leading to skirmishes in connective tissue, the locality of bone and even in the local vasculature. Research in the discipline has uncovered a concerted effort of the microbiome, using an array of survival strategies, to interact with other bacteria and host cells. These strategies aid in colonization by 'ambushing, infiltrating and outflanking' host cells and molecules, responding to local environmental changes (including booby traps for host biomolecules), communicating within and between genera and species that provide MASINT (Measurement and Signature Intelligence) to enhance sustained survival, sabotage the host inflammatory and immune responses and by potentially adopting a 'Fabian strategy' with a war of attrition and resulting disease manifestations. Additionally, much has been learned regarding the ever-increasing complexity of the host-response armamentarium at both cellular and molecular levels that is addressed in this review. Knowledge regarding how these systems fully interact requires both new laboratory and clinical tools, as well as sophisticated modeling of the networks that help maintain homeostasis and are dysregulated in disease. Finally, the triggers resulting in a 'coup de main' by the microbiome (exacerbation of disease) and the characteristics of susceptible hosts that can result in 'pyrrhic victories' with collateral damage to host tissues, the hallmark of periodontitis, remains unclear. While much has been learned, substantial gaps in our understanding of the 'parameters of this war' remain elusive toward fulfilling the Sun Tzu adage: 'If you know the enemy and know yourself, you need not fear the result of a hundred battles.'

The M2 protein from influenza A virus is a 97-amino-acid protein with a single transmembrane helix that forms proton-selective channels essential to virus function. The hydrophobic transmembrane domain of the M2 protein (M2TM) contains a sequence motif that mediates the formation of functional tetramers in membrane environments. A variety of structural models have previously been proposed which differ in the degree of helix tilt, with proposed tilts ranging from ∼15°to 38°. An important issue for understanding the structure of M2TM is the role of peptide-lipid interactions in the stabilization of the lipid bilayer bound tetramer. Here, we labeled the N terminus of M2TM with a nitroxide and studied the tetramer reconstituted into lipid bilayers of different thicknesses using EPR spectroscopy. Analyses of spectral changes provide evidence that the lipid bilayer does influence the conformation. The structural plasticity displayed by M2TM in response to membrane composition may be indicative of functional requirements for conformational change. The various structural models for M2TM proposed to date-each defined by a different set of criteria and in a different environment-might provide snapshots of the distinct conformational states sampled by the protein.Keywords: M2 proton channel; EPR spectroscopy; site-directed spin labeling; membrane protein structure; peptide-lipid interactions; hydrophobic mismatch; helix tilt; lateral pressure The M2 protein from influenza A virus is a 97-amino-acid protein with a single transmembrane helix that forms proton-selective channels essential to virus function. The hydrophobic transmembrane domain of the M2 protein (M2TM) contains a sequence motif that mediates the formation of functional tetramers in membrane environments. Energetics of formation of M2TM have been studied by using analytical ultracentrifugation (Salom et al. 2000;Howard et al. 2002) and thiol-disulfide equilibria (Cristian et al. 2003a,b). A high-resolution crystal structure has not been solved, although a variety of structural models have been proposed based on site-directed mutagenesis in conjunction with computer modeling (Pinto et al. 1997), molecular dynamics calculations (Zhong et al. 2000), infrared spectroscopy (Torres et al. 2000), and solid-state nuclear magnetic resonance spectroscopy (SSNMR) (Kovacs et al. 2000;Wang et al. 2001;Nishimura et al. 2002). The proposed structures are in good agreement with respect to the identities of the side chains lining the pore, the presence of a water-filled pore near the center of the channel, and the packing of monomers with a left-handed tilt. One detail in which models do differ is in the degree of helix tilt, with proposed tilts ranging from approximately 15°to 38°. The 38°angle structure is based on an abundance of high-resolution SSNMR orientational restraints and a single distance restraint (Nishimura et al. 2002), while the 15°angle structure is based on site-directed mutagenesis, and explains a large body of electrophysiological data for this chan...