Studies on the Assembly Characteristics of Large Subunit Ribosomal Proteins in S. cerevisae

During the assembly process of ribosomal subunits, their structural components, the ribosomal RNAs (rRNAs) and the ribosomal proteins (r-proteins) have to join together in a highly dynamic and defined manner to enable the efficient formation of functional ribosomes. In this work, the assembly of large ribosomal subunit (LSU) r-proteins from the eukaryote S. cerevisiae was systematically investigated. Groups of LSU r-proteins with specific assembly characteristics were detected by comparing the protein composition of affinity purified early, middle, late or mature LSU (precursor) particles by semi-quantitative mass spectrometry. The impact of yeast LSU r-proteins rpL25, rpL2, rpL43, and rpL21 on the composition of intermediate to late nuclear LSU precursors was analyzed in more detail. Effects of these proteins on the assembly states of other r-proteins and on the transient LSU precursor association of several ribosome biogenesis factors, including Nog2, Rsa4 and Nop53, are discussed.     

The Genetic Structure and Evolutionary Fate of Parthenogenetic Amphibian Populations as Determined by Markovian Analysis

SYNOPSIS. One-locus, two-allele models are presented which describethe genetic consequences of naturally occurring andexperimentallyinduced parthenogesis in triploid and diploid amphibians. Themodels may in general be used to investigate genetic changeresulting from apomictic (ameiotic) and automictic (meiotic)parthenogenetic reproduction. These models quantify the influence of mutation, segregation,and selection upon genetic variability in parthenogeneticpopulations.They also allow an estimate of the relative importance of stochasticforces in altering this variability. They thus provide a basisfor understanding evolution in these populations. Some of the conclusions derived from this study contradict previouspredictions regarding genetic variability in parthenogeneticpopulations. First, if mutation is the sole source of geneticchange (i.e., strict apomixis), parthenogenetic populationsshould not become completely heterozygous. Second, small amountsof segregation occurring in apomictic populations have enormouseffects upon the genetic variability of these populations, i.e.,they should lose much of their heterozygosity. In addition to these conclusions, the results of this studysuggest that studies of protein variability in parthenogeneticspecies should contribute toward answering the question: Howmuch of the genetic variability observed in nature is evolutionarilyrelevant?     

Dating of graptolite zones by sedimentation rates: implications for rates of evolution

Preliminary determinations of ancient pelagic sedimentation rates agree with modern rates at about 4 meters per million years. By combining data on the thickness of graptolite zones from the North American Cordillera with data from other parts of the world, we have refined the Early Silurian time scale and obtained much better resolution than is possible for radiometric dates. The new Early Silurian time scale allows estimation of true rates of change in graptolite diversity. The Llandoverian diversity explosion is twice as rapid as was previously thought. The brevity of diversity lows and rapidity of speciation support modern theories of quantum evolution.     

In-depth qualitative and quantitative analysis of composite glycosylation profiles and other micro-heterogeneity on intact monoclonal antibodies by high-resolution native mass spectrometry using a modified Orbitrap

Here, we describe a fast, easy-to-use, and sensitive method to profile in-depth structural micro-heterogeneity, including intricate N-glycosylation profiles, of monoclonal antibodies at the native intact protein level by means of mass spectrometry using a recently introduced modified Orbitrap Exactive Plus mass spectrometer. We demonstrate the versatility of our method to probe structural micro-heterogeneity by describing the analysis of three types of molecules: (1) a non-covalently bound IgG4 hinge deleted full-antibody in equilibrium with its half-antibody, (2) IgG4 mutants exhibiting highly complex glycosylation profiles, and (3) antibody-drug conjugates. Using the modified instrument, we obtain baseline separation and accurate mass determination of all different proteoforms that may be induced, for example, by glycosylation, drug loading and partial peptide backbone-truncation. We show that our method can handle highly complex glycosylation profiles, identifying more than 20 different glycoforms per monoclonal antibody preparation and more than 30 proteoforms on a single highly purified antibody. In analyzing antibody-drug conjugates, our method also easily identifies and quantifies more than 15 structurally different proteoforms that may result from the collective differences in drug loading and glycosylation. The method presented here will aid in the comprehensive analytical and functional characterization of protein micro-heterogeneity, which is crucial for successful development and manufacturing of therapeutic antibodies