Secondary Structural Changes in Globular Protein Induced by a Surfactant: Fourier Self-Deconvolution of FT-IR Spectra

Abstract

Conformational changes in ovalbumin, a globular protein, induced by an anionic surfactant, sodium dodecyl sulfate (SDS), have been monitored by an FT-IR spectrometer using ZnSe cylindrical internal reflection optics which allows high quality IR spectra to be obtained in water solution. The most notable change, on addition of SDS, occurs in the composite band of the Amide I absorption band and the vibrational frequency of the composite C=O bond shifts from 1639 cm^?1 to 1652 cm^?1. On the other hand, the position of the Amide II band remains fairly unchanged.

Comparison of the various peak positions in the deconvoluted spectra for the native protein and the perturbed protein clearly shows the effect of SDS on the secondary structures of the protein. SDS unfolds the protein. It increases the helix content slightly. More importantly, it alters the β sheet structure, destroying it almost completely in the Amide I region, while retaining it in its neighbourhood. In the deconvoluted spectra of the perturbed protein, a band at 1531 cm^?1 indicates generation of some β turns. We used the second derivative of the deconvoluted spectra for fixing positions of minor peaks and shoulders.

The results of this study indicate that the deconvolution of the normal IR spectra, consisting of composite bands, provides evidence for the specific secondary structures in a protein and for the way they are affected by changes in the environment, e.g., the addition of SDS. This makes it possible to relate conformational changes to specific secondary structures.     

The PTR family: a new group of peptide transporters

The transport of peptides into cells is a well-documented biological phenomenon which is accomplished by specific, energy-dependent transporters found in a number of organisms as diverse as bacteria and humans. Until recently, the majority of peptide transporters cloned and characterized were found to be proteins of the ATP-binding cassette (ABC) family. We report the identification of a new family of peptide transporters, which we call the PTR family. This group of proteins, distinct from the ABC-type peptide transporters, was uncovered by sequence analyses of a number of recently discovered peptide transport proteins. Alignment of these proteins demonstrated a high number of identical and similar residues and identified conserved glycosylation and phosphorylation sites, as well as a structural motif unique to this group of proteins. Cluster analysis among the proteins indicated these sequences were indeed related and could be further divided into two subfamilies. A phylogenetic analysis of these new peptide transport sequences, compared to over 50 other peptide and membrane-bound transporters, showed that these proteins comprise a distinct, separate group of proteins.     

The aniline blue fluorochrome specifically stains the septum of both live and fixed Schizosaccharomyces pombe cells

Abstract A novel method is described which uses aniline blue for the specific fluorescent staining of the septa of dividing cells of the fission yeast, Schizosaccharomvces pombe . It gives the same results with live and fixed cells. In fixed or, more generally, dead cells there is no staining of the cytoplasm: this renders aniline blue superior to other dyes previously used to stain the septum of S. pombe . This feature allows quantitative analysis of the septum index for fixed samples and, therefore, makes aniline blue the stain of choice for cell cycle kinetic studies.     

The Limited Utility of Multiunit Data in Differentiating Neuronal Population Activity

To date, single neuron recordings remain the gold standard for monitoring the activity of neuronal populations. Since obtaining single neuron recordings is not always possible, high frequency or ‘multiunit activity’ (MUA) is often used as a surrogate. Although MUA recordings allow one to monitor the activity of a large number of neurons, they do not allow identification of specific neuronal subtypes, the knowledge of which is often critical for understanding electrophysiological processes. Here, we explored whether prior knowledge of the single unit waveform of specific neuron types is sufficient to permit the use of MUA to monitor and distinguish differential activity of individual neuron types. We used an experimental and modeling approach to determine if components of the MUA can monitor medium spiny neurons (MSNs) and fast-spiking interneurons (FSIs) in the mouse dorsal striatum. We demonstrate that when well-isolated spikes are recorded, the MUA at frequencies greater than 100Hz is correlated with single unit spiking, highly dependent on the waveform of each neuron type, and accurately reflects the timing and spectral signature of each neuron. However, in the absence of well-isolated spikes (the norm in most MUA recordings), the MUA did not typically contain sufficient information to permit accurate prediction of the respective population activity of MSNs and FSIs. Thus, even under ideal conditions for the MUA to reliably predict the moment-to-moment activity of specific local neuronal ensembles, knowledge of the spike waveform of the underlying neuronal populations is necessary, but not sufficient.     

Brook troot populations in Colorado beaver ponds

Summary A habitat inventory and evaluation of the brook trout populations from 57 beaver ponds in Colorado permitted comparisons of stunted and non-stunted populations of fish and the environmental conditions under which the populations occurred. High standing crops of small fish were usually present in ponds with favorable spawning areas. Seep ponds with few gravel or sandy areas did not favor spawning. As a result, a smaller number of larger fish were recovered. Four-to five-inch stunted trout reached maturity at the end of their second growing season while non-stunted trout matured at the end of their third and fourth growing seasons at lengths of seven to eight inches.     

Phospholipid Composition of Venezuelan Equine Encephalitis Virus

Phospholipid analyses of Venezuelan equine encephalitis virus showed that virus propagated in L-cell monolayers had a higher sphingomyelin content and a lower phosphatidylcholine content than virus grown in chick fibroblast monolayers. Virus of L-cell origin also was found to possess greater thermal stability than virus derived from the chick fibroblast cell.     

Replication and Complementation of Human Adenoviruses and Simian Papovavirus at an Elevated Temperature

The simian papovavirus SV40 replicated as well in simian cells incubated at 41 C as in cells incubated at 37 C, although the latent period was shortened at the elevated temperature. Human adenoviruses differed in their responses to the elevated temperature. Some serotypes, such as 3, 4, 5, 7, 8, 16, and 21, replicated as well, or almost as efficiently, in human cells incubated at 41 C as in cells incubated at 37 C, whereas with other serotypes, such as 1, 2, 6, 12, and 14, maximal yields in cultures incubated at 41 C were much lower than the yields from companion cultures incubated at 37 C. This difference was also detected in simian cells co-infected with SV40 and a human adenovirus; maximal complementation occurred with some serotypes at the elevated temperature but not with other serotypes. The degree of complementation observed in the simian cells at 41 C was directly correlated with the ability of the adenovirus to replicate at 41 C in human cells. Therefore, the capacity of SV40 to serve as a helper virus is not affected by the elevated temperature, showing that the complementation event supplied by the simian virus is heat-stable between 37 and 41 C. Maximal complementation appeared to depend upon a characteristic present in the adenovirus genome.     

Structural analysis of dextrans containing 4-O-α-d-glucosylated α-d-glucopyranosyl residues at the branch points,by use of 13c-nuclear magnetic resonance spectroscopy and gas-liquid chromatography-mass spectrometry

Dextran fractions from NRRL strain Streptococcus sp. B-1526 and the native, structurally homogeneous dextrans from Acetobacter capsulatum B-1225, Leuconostoc mesenteroides B-1307, and L. dextranicum B-1420 were examined by ¹³C-n.m.r. spectroscopy at 90°. Dextran B-1526 fraction I and dextran B-1420 were also examined by g.l.c:-m.s., methylation-structural analysis. All of these dextrans and dextran fractions branch, either primarily or exclusively, through α-d-(1→4)-glucopyranosyl linkages; however, their degrees of branching differ. Several ¹³C-n.m.r. resonances that are diagnostic for 4,6-di-O-substituted α-d-glucopyranosyl residues have been identified. Comparison was made with dextrans from L. mesenteroides B-742 fraction L and Streptobacterium dextranicum B-1254 fraction S[L], for which previously published, methylation-structural analyses had established the presence of 4,6-di-O-substituted α-d-glucopyranosyl residues at the branch points. These fermentation culture, and in a sedimented gum-phase (fraction I). The product from the soluble phase is designated here as fraction S in order to simplify the terminology. Originally⁷, this product was not designated a fraction, because it was, by definition⁸, the main dextran product. The same distinction also applies to the pairs of products from strains B-1380, B-1420, and b-1394 (see ref. 7). The attempts thus made to establish the significance of the phase separation were indeterminant.Methods.— Methods previously described were used for the mythylation⁹ of the dextrans and for structural analysis^6.38 by combined g.l.c-electron-impact mass spectrometry of the aldononitriles. For each permethylation, three successive Hakomori³⁹ methylations were employed on an initial, 40-mg sample, with ～80% (final weight) recovery of each permethylated dextran. Successive formolysis and acetic acid hydrolysis were employed, and, after each step, the resulting solutions were clear, colorless, and free from suspended material. All mass spectra were recorded with a Hewlett-Packard 5980A GC/MS integrated g.l.c.-m.s.-computer system. The g.l.c. peak-integrals reported in Table II were obtained with a Barber-Coleman Series 5000 g.l.c. instrument equipped with hydrogen-flame detectors. On-column injection with glass columns (2mmi.d. x 1.23m) was employed for all chromatograhy.The ¹³C-n.m.r. conditions and the methods for the preparation of dextran samples have been described⁴. In general, a Varian XL-100-15 spectrometer equipped with a Nicolet TT-100 system was employed in the Fourier-transform mode. The dextran samples, ～0.3g/4 mL of deuterium oxide, were maintained at 90°. Chemical shifts are expressed in p.p.m. relative to external tetramethylsilane, but were actually calculated by reference to the solvent lock-signal. The convolution-difference resolution-enhancement (c.d.r.e.) technique has been described⁴⁰.     

Donut-shaped “miniparticles” in nuclei of human and rat cells

Donut-shaped “miniparticles” were extracted from nuclei of various types of human and rat cells. Electron-microscopic investigations showed these particles were predominantly in sucrose density gradient fractions that had an approximate sedimentation coefficient of 21S. These particles were 113±8_A^o in diameter and had an electron dense center of 29±6_A^o. They appeared to be composed of 8 subunits. Quantitative analysis of the number of these particles by electron-micrographic field counting showed nuclei of tumor samples had a larger amount of the particles than the cytosol. However, normal cell cytosol had a larger number of particles than the nuclei. A group of proteins in the 25, 000–33, 000 molecular weight range was shown to be the main protein component by two dimensional gel electrophoresis.