Bovine liver dihydrofolate reductase: purification and properties of the enzyme.

A purification procedure is reported for obtaining bovine liver dihydrofolate reductase in high yield and amounts of 100-200 mg. A key step in the procedure is the use of an affinity gel prepared by coupling pteroyl-L-lysine to Sepharose. The purified reductase has a specific activity of about 100 units/mg and is homogeneous as judged by analytical ultracentrifugation, polyacrylamide gel electrophoresis, and titration with methotrexate. The products of the first step of Edman degradation indicated a minimum purity of 79%. The reductase has a molecular weight of about 21500 on the basis of amino acid composition and 22100 +/- 300 from equilibrium sedimentation. It is not inhibited by antiserum to the Streptococcus faecium reductase (isoenzyme 2). Unlike the reductase of many other vertebrate tissues, the bovine enzyme is inhibited by mercurials rather than activated and it has a single pH optimum at both low and high ionic strength. However, the position of the pH optimum is shifted and the activity increased by increasing ionic strength. Automatic Edman degradation has been used to determine 34 of the amino-terminal 37 amino acid residues. Considerable homology exists between this region and the corresponding regions of the reductase from S. faecium and from Escherichia coli. This strengthens the idea that this region contributes to the structure of the binding site for dihydrofolate.     

Biological vs. physical explanations for the non-random pattern of host occupation by a macroalga attaching to infaunal bivalve molluscs

Summary On a shallow sand flat at Princess Royal Harbour near Albany, Western Australia, the brown macrophyte Hormosira banksii attaches to shells of infaunal bivalves. Hormosira occupies shells of Katelysia rhytiphora in preference to K. scalarina. We proposed and tested four hypotheses to explain this host occupation pattern. First, by following the fate of nearly one thousand marked clams of each species, we rejected the hypothesis that K. rhytiphora exhibits greater longevity and simply possesses more frequent Hormosira because of a longer temporal integration of settlement events. Second, we rejected the hypothesis that K. rhytiphora exhibits higher densities in the top 5 mm of sediment and accumulates more Hormosira because its shell is more abundant in the depth range occupied by the attaching base of Hormosira. Third, we showed that K. rhytiphora because of its larger size is more difficult to dislodge from the sediments than K. scalarina, supporting the hypothesis that Hormosira is rare on K. scalarina because storm waves selectively dislodge and carry to the beach Hormosira attached to K. scalarina. This physical explanation for the Hormosira occupation pattern gets further support from the observation that a third infaunal bivalve, the mussel Brachidontes erosus, has a far higher frequency of Hormosira occupation than either Katelysia species, while providing a much more robust anchor because of its extensive byssal attachments to neighboring mussels. The sizes of Hormosira plants also vary consistently with this physical transport hypothesis: Hormosira is smallest on K. scalarina and largest on B. erosus. Successful colonization of initially unoccupied Katelysia during five 3–9 month periods was also more frequent on K. rhytiphora than on K. scalarina. This suggests a fourth explanation for the Hormosira distribution pattern: that spore settlement is selective for K. rhytiphora in preference to K. scalarina. Although this hypothesis requires further testing, evolution of selective spore settlement would be reasonable given the different likelihoods of subsequent host dislodgement during storm waves.     

Fiber type composition of the plantarflexors of giraffes (Giraffa camelopardalis) at different postnatal stages of development

1. A sample of fibers from deep (close to the bone) and superficial (away from the bone) regions of the plantaris (PLT) and medial (MG) and lateral (LG) gastrocnemius muscles of a neonatal, a 17-day-old and an adult giraffe were typed qualitatively as dark or light based on alkaline preincubation myosin ATPase staining properties and then sized. 2. Each muscle at all ages showed a higher percentage and a larger cross-sectional area (CSA) or light ATPase fibers in the deep than the superficial region. This relationship was qualitatively, although not quantitatively, similar to that reported in hindlimb muscles of other mammals. 3. At all ages, the PLT, the deepest muscle in the synergistic group, had the highest relative total CSA of light ATPase fibers among the muscles sampled. 4. At birth, the PLT had an unusually high percentage of light ATPase fibers in comparison to that found in the same muscle of other mammals. With age, the total CSA of light ATPase fibers increased dramatically in the PLT and decreased slightly in the MG and LG. 5. These data suggest that the PLT, especially the deep portion, may functionally replace the soleus muscle which is absent in the giraffe. In addition, the fiber type results demonstrate that the changes in the fiber type composition of individual muscles observed at different postnatal ages in the giraffe are relatively similar to that reported in smaller mammals, suggesting the existence of similar regulatory mechanisms.     

A pH titration study on the ionic bridging within lipopolysaccharide aggregates

The packing of lipopolysaccharide aggregates from rough strains of Escherichia coli was examined at different pH values. Lipopolysaccharide head-group motion, measured with an electron spin resonance probe, was found to be dependent on pH, and indicated the existence of multiple ionizable groups. Lipopolysaccharide from a rough (Ra) and a heptose-less (Re) mutant were more rigid at pH 5 than at pH 10.5. In addition, head-group mobility of the magnesium salt of Ra lipopolysaccharide was substantially less than that of the sodium salt at pH 7.0, whereas at high pH (pH 12) the two salts were equally fluid. Changes in head-group packing were also reflected in pH-dependent changes in the phase transition measured with differential scanning calorimetry. The enthalpy of the transition, delta Ht, for the sodium salt of Re lipopolysaccharide was greatest at pH 7.5 and approached zero in both the acidic and the basic pH ranges. We propose that fixed charges in the core and lipid A regions significantly influence lipopolysaccharide head-group motion and the lipopolysaccharide aggregation state. Furthermore, ionic bridging among phosphate groups dramatically rigidifies head group interactions in the neutral to acidic pH ranges.