Glycosylated and unglycosylated recombinant-derived human stem cell factors are dimeric and have extensive regular secondary structure.

We have recently described the identification, isolation, and characterization of a factor, termed stem cell factor (SCF), which acts on primitive hematopoietic progenitors of the marrow. A soluble form of the factor was isolated from the conditioned medium of a rat cell line (Zsebo, K. M., Wypych, J., McNiece, I. K., Lu, H. S., Smith, K. A., Karkare, S. B., Sachdev, R. K., Yuschenkoff, V. N., Birkett, N. C., Williams, L. R., Satyagal, V. N., Tung, W., Bosselman, R. A., Mendiaz, E. A., and Langley, K. E. (1990) Cell 63, 195-201) and rat and human cDNAs have been cloned (Martin, F. H., Suggs, S. V., Langley, K. E., Lu, H. S., Ting, J., Okino, K. H., Morris, C. F., McNiece, I. K., Jacobsen, F. W., Mendiaz, E. A., Birkett, N. C., Smith, K. A., Johnson, M. J., Parker, V. P., Flores, J. C., Patel, A. C., Fisher, E. F., Erjavec, H. O., Herrera, C. J., Wypych, J., Sachdev, R. K., Pope, J. A., Leslie, I., Wen, D., Lin, C.-H., Cupples, R. L., and Zsebo, K. M. (1990) Cell 63, 203-211). The cDNAs encode amino acids C-terminal to those found in the isolated natural form, including a putative transmembrane domain. This paper describes the structural characterization of soluble forms of recombinant human SCF purified from Escherichia coli (unglycosylated) and from Chinese hamster ovary (CHO) cells (glycosylated). Fluorescence emission spectra indicate that the single Trp residue is present in a hydrophobic environment. Circular dichroism and infrared spectroscopy indicate considerable secondary structure, including both alpha-helix and beta-sheet. Molecular weight determinations by sedimentation equilibrium show that the molecules are dimeric (noncovalently associated), and gel filtration analyses are consistent with this conclusion. The CHO cell-derived SCF is about 30% carbohydrate by weight, with both N-linked and O-linked sugar. The presence or absence of the carbohydrate does not influence the results of the various structural analyses.     

Mechanism of PKR activation: dimerization and kinase activation in the absence of double-stranded RNA

The kinase PKR is a central component of the interferon antiviral pathway. PKR is activated upon binding double-stranded (ds) RNA to undergo autophosphorylation. Although PKR is known to dimerize, the relationship between dimerization and activation remains unclear. Here, we directly characterize dimerization of PKR in free solution using analytical ultracentrifugation and correlate self-association with autophosphorylation activity. Latent, unphosphorylated PKR exists predominantly as a monomer at protein concentrations below 2 mg/ml. A monomer sedimentation coefficient of s(20,w)(0)=3.58 S and a frictional ratio of f/f(0)=1.62 indicate an asymmetric shape. Sedimentation equilibrium measurements indicate that PKR undergoes a weak, reversible monomer-dimer equilibrium with K(d)=450 microM. This dimerization reaction serves to initiate a previously unrecognized dsRNA-independent autophosphorylation reaction. The resulting activated enzyme is phosphorylated on the two critical threonine residues present in the activation loop and is competent to phosphorylate the physiological substrate, eIF2alpha. Dimer stability is enhanced by approximately 500-fold upon autophosphorylation. We propose a chain reaction model for PKR dsRNA-independent activation where dimerization of latent enzyme followed by intermolecular phosphorylation serves as the initiation step. Subsequent propagation steps likely involve phosphorylation of latent PKR monomers by activated enzyme within high-affinity heterodimers. Our results support a model whereby dsRNA functions by bringing PKR monomers into close proximity in a manner that is analogous to the dimerization of free PKR.     

非Allee哺乳种群系统的最大持续生产量

通过计算和模拟发现：(1)对于给定的净生殖率,非Allee种群系统的最大持续生产量随系统的形状参数的增大而减少;(2)对于给定的形状参数。非Allee种群系统的最大持续生产量随系统的净生殖率的增大而增大。     

Interaction of the trp RNA-binding attenuation protein (TRAP) with anti-TRAP

The trp RNA-binding attenuation protein (TRAP) negatively regulates expression of the tryptophan biosynthesis genes of Bacillus subtilis. In the presence of tryptophan, TRAP is activated to bind to the 5'-leader region of the trp mRNA resulting in termination prior to the structural genes. In addition, accumulation of uncharged tRNA(Trp) induces synthesis of anti-TRAP (AT), which binds to TRAP and inhibits its function. Both of these proteins consist of oligomers of identical subunits. Here, we characterize the self-association of each of these proteins and the TRAP-AT interaction in free solution using equilibrium and velocity analytical ultracentrifugation. TRAP exists as a stable 11-mer in the absence and in the presence of tryptophan. Tryptophan binding induces a conformational change in TRAP. AT exists in a reversible equilibrium between trimer and dodecamer with an equilibrium constant of approximately 3 x 10(14)M(-3). About 20% of the trimer is incompetent to form dodecamer. The AT equilibrium is slow on the time-scale of the velocity experiment. Formation of TRAP-AT complexes occurs only in the presence of tryptophan. A complex containing one TRAP 11-mer and one AT 12-mer forms with high affinity. At higher ratios of TRAP:AT complexes containing two TRAP 11-mers and one AT 12-mer are detected. A model for the structure of the complex is proposed.     

The cerebral proteopathies

The abnormal assembly and deposition of specific proteins in the brain is the probable cause of most neurodegenerative disease afflicting the elderly. These “cerebral proteopathies” include Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), prion diseases, and a variety of other disorders. Evidence is accumulating that the anomalous aggregation of the proteins, and not a loss of protein function, is central to the pathogenesis of these diseases. Thus, therapeutic strategies that reduce the production, accumulation, or polymerization of pathogenic proteins might be applicable to a wide range of some of the most devastating diseases of old age.     

Cerebral Amyloid Angiopathy in an Aged Sooty Mangabey (Cercocebus atys)

A 26-y-old male sooty mangabey (Cercocebus atys) was found at necropsy to have a moderate degree of cerebral amyloid β (Aβ) angiopathy in superficial and parenchymal blood vessels of the brain. Senile (Aβ) plaques were absent, as were neurofibrillary tangles and other signs of neurodegeneration. Affected blood vessels were arterial, capillary, and, less frequently, venous in nature. Histologically, the Aβ40 isoform was more prevalent than was Aβ42. As in humans but unlike in squirrel monkeys, the density of lesions in this mangabey increased along a rostral-to-caudal gradient. Therefore mangabeys appear to conform to the general tendency of nonhuman primates by developing cerebral Aβ angiopathy in the absence of other indices of Alzheimer-type neuropathology.Abbreviations: Aβ, amyloid β, CAA, cerebral amyloid angiopathy, GFAP, glial fibrillary acidic protein, Iba 1, microglia-expressed calcium-binding proteinOne of the most common microvasculopathies in the aging human brain is cerebral amyloid angiopathy (CAA), a disorder in which various aggregation-prone proteins accumulate in the walls of parenchymal and meningeal blood vessels.^4,9 Most often, the amyloidogenic protein is amyloid β (Aβ), a cleavage product of the Aβ precursor protein and the essential component of senile plaques in Alzheimer disease.^13,43 In the brain vasculature, the basal lamina is a primary site of Aβ deposition.^25,35 Severely affected arterioles show a loss of smooth muscle cells in the tunica media, a weakening of the vascular wall and a propensity to rupture.^3,34 CAA thus increases the risk of intracerebral bleeding and may be responsible for as much as 20% of nontraumatic hemorrhagic stroke in elderly humans.^15,18,35 CAA is present to various degrees in virtually all cases of Alzheimer disease,^15,16,21 but it also occurs independently.²⁴ As is the case for other proteopathies, advancing age is a significant risk factor for CAA.^8,19In humans, CAA most often affects the arteries and arterioles of the brain, particularly those in the leptomeninges and cortex.^2,25 CAA is less frequent in veins and capillaries,²⁵ but capillary CAA can be prominent in some cases.^26,33 The occipital lobe is affected most often^1,32,37 but all cortical regions are vulnerable. CAA is variable in occurrence in the cerebellum and uncommon in deep telencephalic gray structures, white matter, and the brainstem,³⁶ except in severely affected cases.³²Although its specific role in the pathogenesis of Alzheimer disease remains uncertain, there is now strong evidence that dementia is exacerbated by CAA.¹⁴ Furthermore, CAA is independently linked to cognitive decline both in rare familial cases²⁰ and in older humans with idiopathic CAA.^2,20 Despite the prevalence of cerebrovascular amyloidosis in elderly humans, surprisingly little is known about its effect on the brain, in part because of a paucity of natural animal models that closely mimic the human disorder.^17,38Nonhuman primates offer a unique opportunity to view CAA from a comparative perspective, given that they normally generate human-sequence Aβ and develop severe cerebral Aβ amyloidosis in old age, generally in the absence of other changes that characterize Alzheimer disease.¹² Nonhuman primates have the additional advantage that, compared with humans, their relatively small brains enable exhaustive regional analysis of microscopic lesions, something that, for practical reasons, is seldom undertaken in the human brain. Here we present the first investigation of age-associated brain changes in sooty mangabeys, focusing in particular on Aβ deposition and related abnormalities. One of the 2 aged mangabeys analyzed had Aβ deposition in the brain which was almost exclusively in the form of CAA. Remarkably, the vessel types affected and the regional distribution of CAA more closely resembled the pattern seen in humans than that in other nonhuman primates, particularly squirrel monkeys.⁶ Differences and similarities in CAA among primate species could provide fresh insights into the development of cerebral amyloidosis and related disorders in older humans.     

Magnetic field measurements near RF induction heaters

We discuss initial magnetic field strength measurements made around radiofrequency (RF) induction heaters. These measurements were made with a monitor developed for the National Institute for Occupational Safety and Health (NIOSH) by the National Bureau of Standards (NBS). The monitor has a dynamic range of .01 to 10,000 A2/m2, a frequency range of 300 kHz to 100 MHz, an isotropic response (+/- .3 dB) with three mutally orthogonal loop antennas that have the ability to measure and display each of three orthogonal magnetic field components, a high probe burnout protection level of 20,000 A2/m2, and an accuracy of +/- 1.0 dB at 13 calibration frequencies. The portable survey monitor was used to measure the magnetic field strengths in the vicinity of 16 RF induction heaters. Typically these induction heaters are operated continuously for several hours. The maximum field strengths (without duty factor correction) ranged from 15 to 4,500 A2/m2 and were measured 5 to 51 cm from the loop applicators of the induction heaters. At locations commonly occupied by workers (ie, approximately 30 to 76 cm from heaters), the fields ranged from .01 to 300 A2/m2 (without duty factor correction).     

Prenatal ionizing irradiaton and early postnatal growth of Colombian and Bolivian squirrel monkeys (Saimiri sciureus)

Fetal squirrel monkeys of Bolivian and Colombian origin were exposed to 100 rad cobalt-60 radiation in utero and carried to term by the dams. Body weight, head circumference, crown–rump length, and food intake were measured during the first 28 postnatal days. Significant subspecies differences were noted in both mothers and offspring, the Colombian squirrel monkeys being generally larger than the Bolivians. The irradiated offspring weighed significantly less, had significantly smaller crania, and consumed significantly less food than control animals. A statistically nonsignificant decrement was also evident in the crown–rump length of radiation-treated infants.