Proteins on ribosome surface: Measurements of protein exposure by hot tritium bombardment technique

The hot tritium bombardment technique [Goldanskii, V. I., Kashirin, I. A., Shishkov, A. V., Baratova, L. A. & Grebenshchikov, N. I. (1988) J. Mol. Biol. 201, 567–574] has been applied to measure the exposure of proteins on the ribosomal surface. The technique is based on replacement of hydrogen by high energy tritium atoms in thin surface layer of macromolecules. Quantitation of tritium radioactivity of each protein has revealed that proteins S1, S4, S5, S7, S18, S20, and S21 of the small subunit, and proteins L7/L12, L9, L10, L11, L16, L17, L24, and L27 of the large subunit are well exposed on the surface of the Escherichia coli 70 S ribosome. Proteins S8, S10, S12, S16, S17, L14, L20, L29, L30, L31, L32, L33, and L34 have virtually no groups exposed on the ribosomal surface. The remaining proteins are found to be exposed to lesser degree than the well exposed ones. No additional ribosomal proteins was exposed upon dissociation of ribosomes into subunits, thus indicating the absence of proteins on intersubunit contacting surfaces.

The emergence of form by replication

It is shown with a simple mathematical model that if a system exhibits a given form (a spatial structure) and is put in contact with another system of the same type but in a state of spatial disorder, then under certain conditions their mutual interaction as they evolve in time allows replication of form in the disordered system with a controllable degree of faithfulness.

Microtubule release from the centrosome

Although microtubules (MTs) are generally thought to originate at the centrosome, a number of cell types have significant populations of MTs with no apparent centrosomal connection. The origin of these noncentrosomal MTs has been unclear. We applied kinetic analysis of MT formation in vivo to establish their mode of origin. Time-lapse fluorescence microscopy demonstrated that noncentrosomal MTs in cultured epithelial cells arise primarily by constitutive nucleation at, and release from, the centrosome. After release, MTs moved away from the centrosome and tended to depolymerize. Laser-marking experiments demonstrated that released MTs moved individually with their plus ends leading, suggesting that they were transported by minus end-directed motors. Released MTs were dynamic. The laser marking experiments demonstrated that plus ends of released MTs grew, paused, or shortened while the minus ends were stable or shortened. Microtubule release may serve two kinds of cellular function. Release and transport could generate the noncentrosomal MT arrays observed in epithelial cells, neurons, and other asymmetric, differentiated cells. Release would also contribute to polymer turnover by exposing MT minus ends, thereby providing additional sites for loss of subunits. The noncentrosomal population of MTs may reflect a steady-state of centrosomal nucleation, release, and dynamics.

Intermediate-term earthquake prediction.

An earthquake of magnitude M and linear source dimension L(M) is preceded within a few years by certain patterns of seismicity in the magnitude range down to about (M - 3) in an area of linear dimension about 5L-10L. Prediction algorithms based on such patterns may allow one to predict approximately 80% of strong earthquakes with alarms occupying altogether 20-30% of the time-space considered. An area of alarm can be narrowed down to 2L-3L when observations include lower magnitudes, down to about (M - 4). In spite of their limited accuracy, such predictions open a possibility to prevent considerable damage. The following findings may provide for further development of prediction methods: (i) long-range correlations in fault system dynamics and accordingly large size of the areas over which different observed fields could be averaged and analyzed jointly, (ii) specific symptoms of an approaching strong earthquake, (iii) the partial similarity of these symptoms worldwide, (iv) the fact that some of them are not Earth specific: we probably encountered in seismicity the symptoms of instability common for a wide class of nonlinear systems.

The use of spreading depression waves for acute and long-term monitoring of the penumbra zone of focal ischemic damage in rats.

Slow potential recording was used for long-term monitoring of the penumbra zone surrounding an ischemic region produced by middle cerebral artery (MCA) occlusion in adult hooded rats (n = 32). Four capillary electrodes (El-E4) were chronically implanted at 2-mm intervals from AP -3, L 2 (El) to AP 0, L 5 (E4). Spontaneous or evoked slow potential waves of spreading depression (SD) were recorded during and 4 h after a 1-h MCA occlusion and at 2- to 3-day intervals afterward for 3 weeks. Duration of the initial focal ischemic depolarization was maximal at E4 and decreased with distance from the focus. SD waves in the penumbra zone were high at El and E2, low and prolonged at E3, and almost absent at E4. Amplitude of elicited SD waves was further reduced 3 days later and slowly increased in the following week. Cortical areas displaying marked reduction of SD waves in the first days after MCA occlusion either remained low or showed substantial (60%) recovery, the probability of which decreased with the duration of the initial focal ischemic depolarization and increased with the distance from the focus. It is concluded that the outcome of ischemia monitored by long-term SD recovery in the perifocal region can be partly predicted from the acute signs of MCA occlusion.

How the first biopolymers could have evolved.

In this work, we discuss a possible origin of the first biopolymers with stable unique structures. We suggest that at the prebiotic stage of evolution, long organic polymers had to be compact to avoid hydrolysis and had to be soluble and thus must not be exceedingly hydrophobic. We present an algorithm that generates such sequences for model proteins. The evolved sequences turn out to have a stable unique structure, into which they quickly fold. This result illustrates the idea that the unique three-dimensional native structures of first biopolymers could have evolved as a side effect of nonspecific physicochemical factors acting at the prebiotic stage of evolution.

Selective pressure of a quinoxaline nonnucleoside inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) on HIV-1 replication results in the emergence of nucleoside RT-inhibitor-specific (RT Leu-74-->Val or Ile and Val-75-->Leu or Ile) HIV-1 mutants.

The quinoxaline nonnucleoside RT inhibitor (NNRTI) (S)-4-isopropoxycarbonyl-6-methoxy-3-(methylthiomethyl)-3,4- dihydroquinoxaline-2(1H)-thione (HBY 097) was used to select for drug-resistant HIV-1 variants in vitro. The viruses first developed mutations affecting the NNRTI-binding pocket, and five of six strains displayed the RT G190-->E substitution, which is characteristic for HIV-1 resistance against quinoxalines. In one variant, a new mutant (G190-->Q) most likely evolved from preexisting G190-->E mutants. The negative charge introduced by the G190-->E substitution was maintained at that site of the pocket by simultaneous selection for V179-->D together with G190-->Q. After continued exposure to the drug, mutations at positions so far known to be specific for resistance against nucleoside RT inhibitors (NRTIs) (L74-->V/I and V75-->L/I) were consistently detected in all cultures. The inhibitory activities of the cellular conversion product of 2',3'-dideoxyinosine (ddI, didanosine), 2',3'-dideoxyadenosine (ddA) and of 2',3'-didehydro-3'-deoxythymidine (d4T, stavudine) against these late-passage viruses were shown to be enhanced with the L74-->V/I RT mutant virus as compared with the wild-type (wt) HIV-1MN isolate. Clonal analysis proved linkage of the codon 74 and codon 75 mutations to the NNRTI-specific mutations in all RT gene fragments. The nonnucleoside- and nucleoside-resistance mutation sites are separated by approximately 35 A. We propose that the two sites "communicate" through the template-primer which is situated in the DNA-binding cleft between these two sites. Quinoxalines cause high selective pressure on HIV-1 replication in vitro; however, the implication of these findings for the treatment of HIV-1 infection has yet to be determined.