The hydrolytic endoribonuclease RNase E, which is widely distributed in bacteria and plants, plays key roles in mRNA degradation and RNA processing in Escherichia coli. The enzymatic activity of RNase E is contained within the conserved amino-terminal half of the 118 kDa protein, and the carboxy-terminal half organizes the RNA degradosome, a multi-enzyme complex that degrades mRNA co-operatively and processes ribosomal and other RNA. The study described herein demonstrates that the carboxy-terminal domain of RNase E has little structure under native conditions and is unlikely to be extensively folded within the degradosome. However, three isolated segments of 10 -40 residues, and a larger fourth segment of 80 residues, are predicted to be regions of increased structural propensity. The larger of these segments appears to be a protein -RNA interaction site while the other segments possibly correspond to sites of self-recognition and interaction with the other degradosome proteins. The carboxy-terminal domain of RNase E may thus act as a flexible tether of the degradosome components. The implications of these and other observations for the organization of the RNA degradosome are discussed.

Neurotrophin receptors corresponding to vertebrate Trk, p75NTR or Sortilin have not been identified in Drosophila, thus it is unknown how neurotrophism may be implemented in insects. Two Drosophila neurotrophins, DNT1 and DNT2, have nervous system functions, but their receptors are unknown. The Toll receptor superfamily has ancient evolutionary origins and a universal function in innate immunity. Here we show that Toll paralogues unrelated to the mammalian neurotrophin receptors function as neurotrophin receptors in fruit-flies. Toll-6 and Toll-7 are expressed in the central nervous system throughout development, and regulate locomotion, motoraxon targeting and neuronal survival. DNT1 and 2 interact genetically with Toll-6 and 7, bind to Toll-7 and 6 promiscuously, and are distributed in vivo in complementary or overlapping domains. We conclude that in fruit-flies, Tolls are not only involved in development and immunity but also in neurotrophism, revealing an unforeseen relationship between the neurotrophin and Toll protein families.

We assess the use to which bioinformatics in the form of bacterial genome sequences, functional gene probes and the protein sequence databases can be applied to hypotheses about obligate autotrophy in eubacteria. Obligate methanotrophy and obligate autotrophy among the chemo- and photo-lithotrophic bacteria lack satisfactory explanation a century or more after their discovery. Various causes of these phenomena have been suggested, which we review in the light of the information currently available. Among these suggestions is the absence in vivo of a functional alpha-ketoglutarate dehydrogenase. The advent of complete and partial genome sequences of diverse autotrophs, methylotrophs and methanotrophs makes it possible to probe the reasons for the absence of activity of this enzyme. We review the role and evolutionary origins of the Krebs cycle in relation to autotrophic metabolism and describe the use of in silico methods to probe the partial and complete genome sequences of a variety of obligate genera for genes encoding the subunits of the alpha-ketoglutarate dehydrogenase complex. Nitrosomonas europaea and Methylococcus capsulatus, which lack the functional enzyme, were found to contain the coding sequences for the E1 and E2 subunits of alpha-ketoglutarate dehydrogenase. Comparing the predicted physicochemical properties of the polypeptides coded by the genes confirmed the putative gene products were similar to the active alpha-ketoglutarate dehydrogenase subunits of heterotrophs. These obligate species are thus genomically competent with respect to this enzyme but are apparently incapable of producing a functional enzyme. Probing of the full and incomplete genomes of some cyanobacterial and methanogenic genera and Aquifex confirms or suggests the absence of the genes for at least one of the three components of the alpha-ketoglutarate dehydrogenase complex in these obligate organisms. It is recognized that absence of a single functional enzyme may not explain obligate autotrophy in all cases and may indeed be only be one of a number of controls that impose obligate metabolism. Availability of more genome sequences from obligate genera will enable assessment of whether obligate autotrophy is due to the absence of genes for a few or many steps in organic compound metabolism. This problem needs the technologies and mindsets of the present generation of molecular microbiologists to resolve it.

Nerve growth factor (NGF) is the ligand for two unrelated cellular receptors, TrkA and p75 NTR , and acts as a mediator in the development and maintenance of the mammalian nervous system. Signaling through TrkA kinase domains promotes neuronal survival, whereas activation of the p75 NTR "death domains" induces apoptosis under correct physiological conditions. However, co-expression of these receptors leads to enhanced neuronal survival upon NGF stimulation, possibly through a ternary p75 NTR ⅐NGF⅐TrkA complex. We have expressed human p75 NTR ligand binding domain as a secreted glycosylated protein in Trichoplusia ni cells. Following assembly and purification of soluble p75 NTR ⅐NGF complexes, mass spectrometry, analytical ultracentrifugation, and solution x-ray scattering measurements are indicative of 2:2 stoichiometry, which implies a symmetric complex. Molecular models of the 2:2 p75 NTR ⅐NGF complex based on these data are not consistent with the further assembly of either symmetric (2:2:2) or asymmetric (2:2:1) ternary p75 NTR ⅐NGF⅐TrkA complexes.Nerve growth factor (NGF) 2 and the structurally and functionally related growth factors (BDNF, neurotrophin (NT) 3 and 4/5) comprise the family of mammalian neurotrophins. In vivo, neurotrophins are secreted as immature progrowth factors, which contain N-terminal propeptides, whose potential function in neurotrophin signaling has been extensively debated recently (for a review, see Ref. 1). The proneurotrophins undergo maturation by cleavage with prohormone convertases, releasing mature neurotrophins with approximate molecular masses of 13 kDa per monomeric unit (for review, see Ref. 2) that are characterized by the cystine knot fold (3). These mature forms are dimeric in solution (ϳ26 kDa).Neurotrophins exert their effects on both neural and non-neural cells through interaction(s) with plasma membrane-bound receptors. In this fashion, they promote survival, differentiation, mitosis, and cell death, depending on the nature and context of the targets and their stage in development (4). They are also unusual among growth factors in that they interact with and activate two distinct classes of receptors that are structurally unrelated. One group, the receptor-tyrosine kinase family (Trk), which has three distinct genes in higher vertebrates encoding the A, B, and C subforms (as well as products of splice variants), shows considerable ligand selectivity, i.e. TrkA binds NGF with high selectivity whereas TrkB binds BDNF and NT 4/5. In contrast, the second type of receptor, the common (or shared) neurotrophin receptor (p75 NTR ), which is a member of the TNF receptor superfamily, binds all of the neurotrophins with about equal affinity (ϳ10 Ϫ9 M) (5). Both types of receptor can and do function in the absence of the other, but there are also many observations that suggest their functions may be linked, perhaps by physical interactions (6). These cross-receptor interactions appear to be particularly important in affecting ligand affinity and in maintaining viability (or n...