Predicting direct protein interactions from affinity purification mass spectrometry data

Background  

Affinity purification followed by mass spectrometry identification (AP-MS) is an increasingly popular approach to observe protein-protein interactions (PPI) in vivo. One drawback of AP-MS, however, is that it is prone to detecting indirect interactions mixed with direct physical interactions. Therefore, the ability to distinguish direct interactions from indirect ones is of much interest.     

Analysis of periodic patterns in amino acid sequences: Collagen

Methods are given for analyzing regularly spaced patterns of amino acids in proteins and applied to the α₁ chain of collagen. Fourier methods use the transform of the sequence either embedded in a very long array or folded onto a fundamental base period. Filtering through a moveable “window” of definite width is used to display almost regular features at any chosen frequency. A pattern detection method is described for patterns of general shape. Collagen has statistically significant periodicities at fractions of the stagger distance D = 670 Å. Hydrophobic groups show strong orders of 5, 6, 11; proline 5; charged groups 6, 18, 21. Charged residues mostly occur as neutral pairs. Their distribution has strong 6th and 21st orders which also appear in the changes which are paired at multiples of D. Charge pairs separated by (D + 3) residues show a strong 5D/89 pattern and may form a system of salt bridges across the fibril. There is no sign of any regular pattern of amino acids over the triple helix with a period close to its natural pitch of 30 residues. Supercoiled models with six relative turns of the contact edge between paired triple-helical strands are examined.     

Linker histone-DNA complexes: enhanced stability in the presence of aluminum lactate and implications for Alzheimer's disease

The binding of human brain linker histone proteins to a radiolabelled human Alu repetitive element was examined by mobility shift assay. Analysis of the complexes formed from protein extracts of whole neocortical nuclei, under physiological conditions in vitro revealed that linker histone H1(0) has the highest affinity for the Alu DNA sequence. The linker histone-DNA complexes assembled in the presence of aluminum lactate were more resistant to sodium chloride-induced dissociation than those formed in the presence of sodium lactate. The enhanced stability of deoxyribonucleoprotein (DNP) complexes in the presence of the aluminum cation may be of significance in neurodegenerative conditions such as Alzheimer's disease where aluminum preferentially associates with DNA containing structures of the nucleus.     

The effect of cancer on the spin-lattice relaxation time of mouse blood and tissue

Spin-lattice relaxation time measurements were made on the muscle, heart, kidneys, spleen, liver, brain and blood of both normal mice and those injected with Sarcoma-180. As well as a marked increase in the relaxation time of the tumout itself, mice injected with Sarcoma-180 showed a rise in the kidney relaxation time, along with blood relaxation times which were often either above or below the range found in normal mice.Correlation coefficients for the relaxation times of the various organs were calculated and found to be very sensitive to the physiological state of the animal. For healthy mice, the correlatiosn were reasonably high, but even mild stress decreased the correlated, whilst the presence of cancer almost completely destroyed the correlations. These findings are all consistent with the hypothesis that cancer affects the water regulatory system of the animal as a whole.     

A microwell assay for anchorage independent cell growth

We describe modifications of the conventional assay for anchorage independent growth of fibroblasts that enable the assay to be carried out in microwell plates, as opposed to the conventional Petri dishes. The microwell assay is a good discriminator of final EGF concentrations in the range 10-100 picograms/ml, and can be used to detect absolute amounts of EGF below 2 pg. Addition of TGF beta to EGF enhances colony formation in the microassay in the usual manner. We describe the use of this microassay to identify and map local production of transforming growth factor activity by the component pieces of individual chick embryos. Transforming activity was identified in all the stages tested (Hamburger and Hamilton stages 13-23). Highest levels were found near the mid-line of the embryo. No clear differences in the cranio-caudal axis have so far been identified. This technique will enable the spatial and temporal distribution of transforming activity throughout vertebrate embryos to be completely mapped. It seems likely that this mapping process will help elucidate the normal role of transforming growth factors in embryos.     

Effects of low temperature on the burrowing rates of four sandy beach molluscs

Burrowing rates have been measured as a function of size and temperature in four molluscs from warm temperate South African sandy beaches: two bivalves of the genus Donax and two whelks of the genus Bullia. Animals were acclimated to a summer temperature of 21 °C and then subjected to drastic temperature drops, aimed at simulating coastal temperatures recorded during local upwelling. Burrowing times increased from 6–16 s at 20 °C to 18-∞ s at 10 °C, the lowest temperature recorded during summer upwelling. The degree of retardation of burrowing that occurred at low temperature was inversely proportional to the height of penetration into the intertidal zone by the different species. Large animals burrowed more slowly than small animals but were less affected by lowered temperature. Comparison of burrowing rates at 10 °C against swash frequencies recorded under upwelling conditions suggests that under extreme conditions of upwelling, burrowing in three species may not be rapid enough to prevent some individuals being swept up the beach by the swash. The most dangerous time in this respect is at the high tide during choppy sea conditions.     

The evolutionary ecology of detritus feeding in the larvae of freshwater Diptera

Detritus (dead organic matter), largely of terrestrial origin, is superabundant in inland waters but because of its indigestible nature, would appear to be a poor food source for animals. Yet this unpromising material is widely used as food and indeed can be viewed as a defining characteristic of the freshwater environment. We here explore the relationships among animals, detritus and its associated micro-organism decomposers, taking a functional approach. We pose questions about interrelationships and attempt to arrive at new insights by disentangling them from an adaptive point of view. To do this we have been careful in selecting the habitats for detailed consideration. Rain pools on rock surfaces in tropical Africa and pools on peat moorland in the UK were chosen. Both examples have a relatively simple community structure and hence offer the prospect of achieving our aim. As model organisms for study we focus principally on the aquatic stages of selected holometabolous insects; that is, selected genera of the universally common midges, Ceratopogonidae and Chironomidae. We approach these case studies from an evolutionary ecology perspective and see detritus as a simple template upon which a beautiful complex of adaptations can evolve.     

The computation of simulated endplate potentials

A method is described for computing simulated subthreshold responses (.e. endplate potentials) for the Falk-Fatt (1964) cable model of a muscle fibre due to a punctate change in ionic conductance. The method has been applied to a comparison of the Falk-Fatt and classical models. Comparison with experimental results suggests that the model is adequate to account for the response of the mouse muscle fibre, in which the endplate is highly localized. For frog muscle there are larger discrepancies, which may be due to the extended endplate in this species.