A Group Sequential Method for one Standard Control and more than one Experimental Treatment

Many group-sequential test procedures have been proposed to meet the ethical need for interim analyses. All of these papers, however, focus their discussion on the situation where there are only one standard control and one experimental treatment. In this paper, we consider a trial with one standard control, but with more than one experimental treatment. We have developed a group-sequential test procedure to accommodate any finite number of experimental treatments. To facilitate the practical application of the proposed test procedure, on the basis of Monte Carlo simulation, we have derived the critical values of α-levels equal to 0.01, 0.05 and 0.10 for the number of experimental treatments ranging from 2 to 4 and the number of multiple group sequential analysis ranging from 1 to 10. Comparing with a single non-sequential analysis that has a reasonable power (say, 0.80), we have demonstrated that the application of the proposed test procedure may substantially reduce the required sample size without seriously sacrificing the original power.     

MPN tables for more than one test

Summary MPN tables are presented, restricted to those tube combinations that are acceptable considering the number of tests performed (1, 2, 3, 5, or 10). Confidence limits of 95 and 99% are given for each result.     

Virus membrane-fusion proteins: more than one way to make a hairpin

Structure-function studies have defined two classes of viral membrane-fusion proteins that have radically different architectures but adopt a similar overall 'hairpin' conformation to induce fusion of the viral and cellular membranes and therefore initiate infection. In both classes, the hairpin conformation is achieved after a conformational change is triggered by interaction with the target cell. This review will focus in particular on the properties of the more recently described class II proteins.     

The sulfhydryl groups of the thiol-dependent cytolytic toxin from Bacillus alvei evidence for one essential sulfhydryl group

Alveolysin, an extracellular protein toxin (M_r ? 63,000) excreted by Bacillus alvei and purified to homogeneity was shown to contain four cysteine residues. All thiol groups of the hemolytically active toxin preparation were free as found by direct titration by 5,5′-dithiobis (2-nitrobenzoic acid) and confirmed by the absence of disulfide bond. Toxin alkylation with tosyl lysine chloromethyl ketone resulted in the complete loss of hemolytic activity and the disappearance of only one thiol group with no modification of histidine residues. These results support the conclusion that one essential thiol group is implicated in the membrane-disrupting activity of alveolysin.     

Hydrogen bonds and masking in the sulfhydryl group in proteins

Szent-Györgi has shown that the relative amounts of readily titratable (“free”) SH groups vs. those readily titratable only following denaturation (“masked”) varies significantly from normal to cancerous organ tissues. It is therefore important to inquire into the nature of the two forms of protein-borne SH. Of the four suggested mechanisms for the “masking” of protein SH groups toward hydrophilic reagents, namely: (i) sequestration in hydrophobic regions—whether between chain-folds or between agglomerated protein sub-units, (ii) local steric hindrance, (iii) cyclic hydrogen-bonding to local peptide amino acid residues, or (iv) covalent bonding as in thiazoldines; the first mechanism, that of sequestration in hydrophobic regions appears from present evidence to be the most likely cause. Various spectroscopic, reaction rate and entropy arguments are presented and compared which lead to this conclusion.In addition we have calculated the binding energy of the SH group to various sets of lone pair electrons appropriate to N, O, F, P, S, and Cl atoms in molecules. The calculation was made in the configuration interaction of valence orbitals (CIVO) scheme and gave binding energies and interatomic distances in reasonable agreement with available experimental data.     

RGS proteins: more than just GAPs for heterotrimeric G proteins

Members of the newly described RGS family of proteins have a common RGS domain that contains GTPase-activating activity for many Galpha subunits of heterotrimeric G proteins. Their ability to dampen signalling via Galphai-, Galphaq- and Galpha12/13-coupled pathways makes them crucial players in mediating the multitude of cellular processes controlled by heterotrimeric G proteins. Some RGS proteins also contain additional motifs that link them to other signalling networks, where they constitute effector-type molecules. This review summarizes recent findings on RGS proteins, especially those that implicate RGS proteins in more than just enhancing the GTPase activity of their Galpha subunit targets.     

Model for microbial growth with more than one limiting nutrient

Microbial growth rate is determined by concentrations of nutrients. Our new model uses conventional Monod formulations and weighs the contribution of potentially limiting nutrients according to their half-saturation constants. The model fits our data and that of others better than do older models, even though no new coefficients were added.     

Proteomic analysis of a pleistocene mammoth femur reveals more than one hundred ancient bone proteins

We used high-sensitivity, high-resolution tandem mass spectrometry to shotgun sequence ancient protein remains extracted from a 43?000 year old woolly mammoth ( Mammuthus primigenius ) bone preserved in the Siberian permafrost. For the first time, 126 unique protein accessions, mostly low-abundance extracellular matrix and plasma proteins, were confidently identified by solid molecular evidence. Among the best characterized was the carrier protein serum albumin, presenting two single amino acid substitutions compared to extant African ( Loxodonta africana ) and Indian ( Elephas maximus ) elephants. Strong evidence was observed of amino acid modifications due to post-mortem hydrolytic and oxidative damage. A consistent subset of this permafrost bone proteome was also identified in more recent Columbian mammoth ( Mammuthus columbi ) samples from temperate latitudes, extending the potential of the approach described beyond subpolar environments. Mass spectrometry-based ancient protein sequencing offers new perspectives for future molecular phylogenetic inference and physiological studies on samples not amenable to ancient DNA investigation. This approach therefore represents a further step into the ongoing integration of different high-throughput technologies for identification of ancient biomolecules, unleashing the field of paleoproteomics.     

Evidence for more than one pathway in the formation of purine deoxyribonucleotides

The biosynthesis of purine deoxyribonucleotides was studied in the context of general purine nucleotide metabolism in the chick with the aid of radioactive nucleic acid precursors. Our results showed that in chick liver and intestine, the nucleoside phosphate reductase system so firmly established in E. coli [1] and L. leichmanni [2] is not exclusively responsible for the biosynthesis of purine deoxyribonucleotides.     

F-box proteins: more than baits for the SCF?

Progression through the mammalian cell cycle is associated with the activity of four cyclin dependent kinases (Cdc2/Cdk1, Cdk2, Cdk4, and Cdk6). Knockout mouse models have provided insight into the interplay of these Cdks. Most of these models do not exhibit major cell cycle defects revealing redundancies, and suggesting that a single Cdk might be sufficient to drive the cell cycle, similar as in yeast. Recent work on Cdk2/Cdk4 double knockouts has indicated that these two Cdks are required to phosphorylate Rb during late embryogenesis. The lack of Rb phosphorylation is progressive and associated with reduced E2F-inducible gene expression. Cdk2 and Cdk4 share the essential function of coupling the G1/S transition with mitosis. However, proliferation in early embryogenesis appears to be independent of Cdk2 and Cdk4. We discuss these observations and propose molecular mechanisms that establish the requirement for Cdk2 and Cdk4 at the G1/S transition. We are considering that the balance between proliferation and differentiation is disturbed, which affects especially heart development and leads to embryonic lethality in Cdk2 ^-/- Cdk4 ^-/- mutants. We also discuss the specific functions of Cdk4 and Cdk6, which ironically do not compensate for each other.     

Evidence for more than one Ca2+ transport mechanism in mitochondria.

The active transport and internal binding of the Ca2+ analogue Mn2+ by rat liver mitochondria were monitored with electron paramagnetic resonance. The binding of transported Mn2+ depended strongly on internal pH over the range 7.7-8.9. Gradients of free Mn2+ were compared with K+ gradients measured on valinomycin-treated samples. In the steady state, the electrochemical Mn2+ activity was larger outside than inside the mitochondria. The observed gradients of free Mn2+ and of H+ could not be explained by a single "passive" uniport or antiport mechanism of divalent cation transport. This conclusion was further substantiated by observed changes in steady-state Ca2+ and Mn2+ distributions induced by La3+ and ruthenium red. Ruthenium red reduced total Ca2+ or Mn2+ uptake, and both inhibitors caused release of divalent cation from preloaded mitochondria. A model is proposed in which divalent cations are transported by at least two mechanisms: (1) a passive uniport and (2) and active pump, cation antiport or anion symport. The former is more sensitive to La3+ and ruthenium red. Under energized steady-state conditions, the net flux of Ca2+ or Mn2+ is inward over (1) and outward over (2). The need for more than one transport system inregulating cytoplasmic Ca2+ is discussed.     

Evidence for more than one mechanism of action of the glucocorticoid hormones

The data presented here provide evidence for at least two major cellular responses to the glucocorticoid hormones that do not seem to involve an interaction between the activated hormone-receptor complex and chromatin. We observed that 5mM pyridoxine in L-cell culture medium both increases the intracellular content of free pyridoxal phosphate by about 30% and decreases the glucocorticoid induction of glutamine synthetase to less than 30% of that at the normal (5 microM) concentration of the vitamin. Two other responses to the hormone-a major decrease in the polysome content of the cells and a decrease in the state of phosphorylation of ribosomal protein S6-are unaffected by the 5mM dose of the vitamin. In light of the findings of DiSorbo and Litwack ((1981) Biochem. Biophys. Res. Commun. 99, 1203-1208), pyridoxal phosphate appears to modulate the enzyme-inducing activity of dexamethasone in L-cells but not the polysome and ribosome related changes. These results are consistent with the hypothesis that certain important glucocorticoid induced responses are independent of nuclear function.     

Evidence for more than one Parkinson's disease-associated variant within the HLA region

Parkinson''s disease (PD) was recently found to be associated with HLA in a genome-wide association study (GWAS). Follow-up GWAS''s replicated the PD-HLA association but their top hits differ. Do the different hits tag the same locus or is there more than one PD-associated variant within HLA? We show that the top GWAS hits are not correlated with each other (0.00≤r²≤0.15). Using our GWAS (2000 cases, 1986 controls) we conducted step-wise conditional analysis on 107 SNPs with P<10⁻³ for PD-association; 103 dropped-out, four remained significant. Each SNP, when conditioned on the other three, yielded P_SNP1 = 5×10⁻⁴, P_SNP2 = 5×10⁻⁴, P_SNP3 = 4×10⁻³ and P_SNP4 = 0.025. The four SNPs were not correlated (0.01≤r²≤0.20). Haplotype analysis (excluding rare SNP2) revealed increasing PD risk with increasing risk alleles from OR = 1.27, P = 5×10⁻³ for one risk allele to OR = 1.65, P = 4×10⁻⁸ for three. Using additional 843 cases and 856 controls we replicated the independent effects of SNP1 (P_{conditioned-on-SNP4} = 0.04) and SNP4 (P_{conditioned-on-SNP1} = 0.04); SNP2 and SNP3 could not be replicated. In pooled GWAS and replication, SNP1 had OR_{conditioned-on-SNP4} = 1.23, P_{conditioned-on-SNP4} = 6×10⁻⁷; SNP4 had OR_{conditioned-on-SNP1} = 1.18, P_{conditioned-on-SNP1} = 3×10⁻³; and the haplotype with both risk alleles had OR = 1.48, P = 2×10⁻¹². Genotypic OR increased with the number of risk alleles an individual possessed up to OR = 1.94, P = 2×10⁻¹¹ for individuals who were homozygous for the risk allele at both SNP1 and SNP4. SNP1 is a variant in HLA-DRA and is associated with HLA-DRA, DRB5 and DQA2 gene expression. SNP4 is correlated (r² = 0.95) with variants that are associated with HLA-DQA2 expression, and with the top HLA SNP from the IPDGC GWAS (r² = 0.60). Our findings suggest more than one PD-HLA association; either different alleles of the same gene, or separate loci.