Fibril modelling by sequence and structure conservation analysis combined with protein docking techniques: β2-microglobulin amyloidosis

Obtaining atomic resolution structural models of amyloid fibrils is currently impossible, yet crucial for our understanding of the amyloid mechanism. Different pathways in the transformation of a native globular domain to an amyloid fibril invariably involve domain destabilization. Hence, locating the unstable segments of a domain is important for understanding its amyloidogenic transformation and possibly control it. Since relative conservation is suggested to relate to local stability [H. Benyamini, K. Gunasekaran, H. Wolfson, R. Nussinov, Conservation and amyloid formation: a study of the gelsolin-like family, Proteins 51 (2003) 266–282. [24]], we performed an extensive, sequence and structure conservation analysis of the β₂-microglobulin (β₂-m) domain. Our dataset include 51 high resolution structures belonging to the “C1 set domain” family and 132 clustered PSI-BLAST search results. Segments of the β₂-m domain corresponding to strands A (residues 12–18), D (45–55) and G (91–95) were found to be less conserved and stable, while the central strands B (residues 22–28), C (36–41), E (62–70) and F (78–83) were found conserved and stable. Our findings are supported by accumulating observations from various experimental methods, including urea denaturation, limited proteolysis, H/D exchange and structure determination by both NMR and X-ray crystallography. We used our conservation findings together with experimental literature information to suggest a structural model for the polymerized unit of β₂-m. Pairwise protein docking and subsequent monomer stacking in the same manner suggest a fibril model consistent with the cross-β structure.     

Perimenstrual behavior changes among female yellow baboons: Some similarities to premenstrual syndrome (PMS) in women

Although numerous studies have documented changes in the behavior of nonhuman primate females around the time of ovulation, very little attention has been paid to behavior changes around the time of menstruation. Yet such information is obviously relevant to under standing the origins and etiology of the adverse mood and behavior changes experienced premenstrually by many women. Analysis of data obtained during 115 hours of observation on 13 female Amboseli (Kenya) baboons (Papio cynocephalus) during 24 menstrual periods showed that prior to and during the time of menstrual onset, these individuals exhibited distinct changes in their activity budgets, nearest-neighbor distances, and patterns of social interaction. Furthermore, in comparison to females around the time of ovulation, perimenstrual females showed increased rates of agonistic interaction and decreased (but nonzero) rates of sexual interaction with adult males. These premenstrual and perimenstrual behavior changes among female yellow baboons thus show some intriguing similarities to several commonly reported behavioral symptoms of premenstrual syndrome (PMS) in women.     

E46K Parkinson’s-Linked Mutation Enhances C-Terminal-to-N-Terminal Contacts in α-Synuclein

Parkinson's disease (PD) is associated with the deposition of fibrillar aggregates of the protein α-synuclein (αS) in neurons. Intramolecular contacts between the acidic C-terminal tail of αS and its N-terminal region have been proposed to regulate αS aggregation, and two originally described PD mutations, A30P and A53T, reportedly reduce such contacts. We find that the most recently discovered PD-linked αS mutation E46K, which also accelerates the aggregation of the protein, does not interfere with C-terminal-to-N-terminal contacts and instead enhances such contacts. Furthermore, we do not observe a substantial reduction in such contacts in the two previously characterized mutants. Our results suggest that C-terminal-to-N-terminal contacts in αS are not strongly protective against aggregation, and that the dominant mechanism by which PD-linked mutations facilitate αS aggregation may be altering the physicochemical properties of the protein such as net charge (E46K) and secondary structure propensity (A30P and A53T).     

Effects of dibutyryl cyclic AMP on tyrosine uptake and metabolism in neuroblastoma cultures

A series of thin-layer Chromatographic (TLC) systems were employed to study the effects of dibutyryl cyclic AMP (db-cAMP) on the metabolism of ³H-tyrosine in neuroblastoma cultures. The neuroblastoma monolayer cultures incubated with radiolabelled tyrosine synthesized di-hydroxyphenylalanine (DOPA), dopamine (DA), and norepinephrine (NE), in confirmation of previous reports identifying these compounds in neuroblastoma cultures. In addition, we found evidence suggesting the presence of metabolites of DA and NE, that is, homovanillic acid (HVA) and 3-methoxy-4-hydroxyphenylglycol (MHPG) together with 3-methoxy-4-hydroxymandelic acid (VMA). When these cultures were grown in the presence of db-cAMP for 3 days, tyrosine uptake was increased with a proportional increase in tyrosine hydroxylation. This effect persisted in the absence of db-cAMP, but it was not apparent with only 90 min exposure to db-cAMP. Suspension cultures showed the same baseline level of tyrosine uptake as did monolayer cultures, but the uptake in suspension cultures failed to increase with db-cAMP treatment. It is suggested that the db-cAMP induced differentiation of the neuroblastoma cells in monolayer cultures was associated with induction of a tyrosine uptake system.     

Sweet corn phosphorylase: purification and properties

Sweet corn 1,4-α-glucan phosphorylase was purified 190-fold to a near homogeneous state. The enzyme had a molecular weight of about 315,000 on Sephadex G-200 chromatography. The pyridoxal 5′-phosphate content was found to 1 mole per 140,000 g protein, suggesting that the enzyme is dimeric. On sucrose density gradient ultracentrifugation the sweet corn phosphorylase was dissociated to an active monomeric species with a molecular weight of 150,000 and a sedimentation coefficient of 8 S. The priming specificity of the sweet corn phosphorylase was investigated; maltose was not a primer and maltotriose was the smallest apparent primer. The Michaelis constants for the maltosaccharide series from maltopentaose to maltooctaose were determined. The effect of d-enzyme on the apparent priming specificity of the enzyme was investigated. Adenosine diphosphoglucose and 2,3-diphosphoglycerate were found to inhibit the enzyme activity.     

Letter: Myosin filaments showing a 430 A axial repeat periodicity

Synthetic myosin filaments with regular projections at intervals of 430 Å along their entire lengths have been observed using the electron microscope. The filaments were formed following dialysis in or rapid dilution to 0.3 m-KC1, 0.01 m-imidazole or phosphate buffer, pH 7.0. It is suggested that these filaments are made up of myosin molecules staggered by 430 Å.     

Differentiation of a human monocyte-like cell line by (2′–5′) oligoisoadenylate

Treatment of a human monocyte-like cell line (U-937) by (2'-5')ApApA, the 5' dephosphorylated product of (2'-5')oligo-isoadenylate [oligo(A)] synthetase, an interferon-induced enzyme, was able to induce differentiation, mimicking the effect of interferon treatment. Treatment of U-937 cells with (2'-5')ApApA resulted in morphologic changes, new (monocyte-associated) membrane antigen expression, and acquisition of the capacity to mediate antibody-dependent cellular cytotoxicity (ADCC). (2'-5')ApA and (3'-5')ApApA were without effect. A myeloid cell line (HL-60) which differentiates in response to other agents, but not to alpha-interferon, was not able to differentiate in response to (2'-5')ApApA, despite the ability of interferon to induce (2'-5')oligo (A) synthetase.