Ubiquitin-specific proteases of Saccharomyces cerevisiae. Cloning of UBP2 and UBP3, and functional analysis of the UBP gene family.

In eukaryotes, both natural and engineered ubiquitin (Ub) fusions to itself or other proteins are cleaved by processing proteases after the last (Gly76) residue of ubiquitin. YUH1 and UBP1, the genes for two ubiquitin-specific proteases of the yeast Saccharomyces cerevisiae, have been cloned previously and shown to encode nonhomologous proteins. Using an Escherichia coli-based genetic screen, we have isolated two other yeast genes for ubiquitin-specific proteases, named UBP2 and UBP3. Ubp2 (1,264 residues), Ubp3 (912 residues), and the previously cloned Ubp1 (809 residues) are largely dissimilar except for two short regions containing Cys and His which encompass their putative active sites. Neither of these proteases has sequence similarities to Yuh1. Both Ubp2 and the previously identified Ubp1 cleave in vitro at the C terminus of the ubiquitin moiety in natural and engineered fusions irrespective of their size, poly-Ub being the exception. However, both Ubp1 and Ubp2 are also capable of cleaving poly-Ub when coexpressed with it in E. coli, suggesting that such cleavage is largely cotranslational. Although inactive in E. coli extracts, Ubp3 was active with all of the tested ubiquitin fusions except poly-Ub when coexpressed with them in E. coli. Null yuh1 ubp1 ubp2 ubp3 quadruple mutants are viable and retain the ability to deubiquitinate ubiquitin fusions, indicating the presence of at least one more ubiquitin-specific processing protease in S. cerevisiae.     

Nanosecond time scale folding dynamics of a pentapeptide in water

Reverse turns, four-residue sections of polypeptides where the chain changes direction by about 180 degrees, are thought to be important protein folding initiation structures. However, the time scale and mechanism for their formation have yet to be determined experimentally. To develop a microscopic picture of the formation of protein folding initiation structures, we have carried out a pair of 2.2-ns molecular dynamics simulations of Tyr-Pro-Gly-Asp-Val, a peptide which is known to form a high population of reverse turns in water. In the first simulation, which was started with the peptide in an ideal type II reverse turn involving the first four residues, the turn unfolded after about 1.4 ns. After about 0.6 ns in the second simulation, which was started with the peptide in a fully extended conformation, the peptide folded into a type II turn which had a transient existence before unfolding. The peptide remained unfolded for another 0.9 ns before folding into a type I turn involving the last four residues. The type I turn lasted for about 0.2 ns before unfolding. Thus, these simulations showed that protein folding initiation structures can form and dissolve on the nanosecond time scale. Furthermore, the atomic-level detail of the simulations allowed us to identify some of the interactions which can stabilize the folded structures. The type II turns were stabilized by either a salt bridge between the terminal groups or a backbone-C-terminal group hydrogen bond, and the type I turns were stabilized by a hydrophobic interaction between the proline and valine-side chains.     

A large‐area,spatially continuous assessment of land cover map error and its impact on downstream analyses

Land cover maps increasingly underlie research into socioeconomic and environmental patterns and processes, including global change. It is known that map errors impact our understanding of these phenomena, but quantifying these impacts is difficult because many areas lack adequate reference data. We used a highly accurate, high‐resolution map of South African cropland to assess (1) the magnitude of error in several current generation land cover maps, and (2) how these errors propagate in downstream studies. We first quantified pixel‐wise errors in the cropland classes of four widely used land cover maps at resolutions ranging from 1 to 100 km, and then calculated errors in several representative “downstream” (map‐based) analyses, including assessments of vegetative carbon stocks, evapotranspiration, crop production, and household food security. We also evaluated maps’ spatial accuracy based on how precisely they could be used to locate specific landscape features. We found that cropland maps can have substantial biases and poor accuracy at all resolutions (e.g., at 1 km resolution, up to ～45% underestimates of cropland (bias) and nearly 50% mean absolute error (MAE, describing accuracy); at 100 km, up to 15% underestimates and nearly 20% MAE). National‐scale maps derived from higher‐resolution imagery were most accurate, followed by multi‐map fusion products. Constraining mapped values to match survey statistics may be effective at minimizing bias (provided the statistics are accurate). Errors in downstream analyses could be substantially amplified or muted, depending on the values ascribed to cropland‐adjacent covers (e.g., with forest as adjacent cover, carbon map error was 200%–500% greater than in input cropland maps, but ～40% less for sparse cover types). The average locational error was 6 km (600%). These findings provide deeper insight into the causes and potential consequences of land cover map error, and suggest several recommendations for land cover map users.     

<Emphasis Type="Italic">Rht24</Emphasis> reduces height in the winter wheat population ‘Solitär × Bussard’ without adverse effects on Fusarium head blight infection

Key message

The dwarfing gene Rht24 on chromosome 6A acts in the wheat population ‘Solitär × Bussard’, considerably reducing plant height without increasing Fusarium head blight severity and delaying heading stage.

Abstract

The introduction of the Reduced height (Rht)-B1 and Rht-D1 semi-dwarfing genes led to remarkable increases in wheat yields during the Green Revolution. However, their utilization also brings about some unwanted characteristics, including the increased susceptibility to Fusarium head blight. Thus, Rht loci that hold the potential to reduce plant height in wheat without concomitantly increasing Fusarium head blight (FHB) susceptibility are urgently required. The biparental population ‘Solitär × Bussard’ fixed for the Rht-1 wild-type alleles, but segregating for the recently described gibberellic acid (GA)-sensitive Rht24 gene, was analyzed to identify quantitative trait loci (QTL) for FHB severity, plant height, and heading date and to evaluate the effect of the Rht24 locus on these traits. The most prominent QTL was Rht24 on chromosome 6A explaining 51% of genotypic variation for plant height and exerting an additive effect of ? 4.80 cm. For FHB severity three QTL were detected, whereas five and six QTL were found for plant height and heading date, respectively. No FHB resistance QTL was co-localized with QTL for plant height. Unlike the Rht-1 semi-dwarfing alleles, Rht24b did not significantly affect FHB severity. This demonstrates that the choice of semi-dwarfing genes used in plant breeding programs is of utmost consideration where resistance to FHB is an important breeding target.

     

Protein‐engineering of chitosanase from Bacillus sp. MN to alter its substrate specificity

Partially acetylated chitosan oligosaccharides (paCOS) have various potential applications in agriculture, biomedicine, and pharmaceutics due to their suitable bioactivities. One method to produce paCOS is partial chemical hydrolysis of chitosan polymers, but that leads to poorly defined mixtures of oligosaccharides. However, the effective production of defined paCOS is crucial for fundamental research and for developing applications. A more promising approach is enzymatic depolymerization of chitosan using chitinases or chitosanases, as the substrate specificity of the enzyme determines the composition of the oligomeric products. Protein‐engineering of these enzymes to alter their substrate specificity can overcome the limitations associated with naturally occurring enzymes and expand the spectrum of specific paCOS that can be produced. Here, engineering the substrate specificity of Bacillus sp. MN chitosanase is described for the first time. Two muteins with active site substitutions can accept N‐acetyl‐D‐glucosamine units at their subsite (?2), which is impossible for the wildtype enzyme.     

Immunological and functional characterization of Mycobacterium leprae protein antigens: an overview

A major focus of leprosy research in the last 10 years has been the identification and characterization of antigens of Mycobacterium leprae that interact with antibodies and T cells of the host's immune response. Through the combined efforts of many different laboratories, a substantial number of protein antigens have been identified and characterized. In this MicroReview we present an updated list of M. leprae protein antigens, and, with emphasis on recent developments, summarize what is known regarding their functional and immunological features.     

Side-chain conformation and dynamics in a solid peptide: CP-MAS NMR study of valine rotamers and methyl-group relaxation in fully 13C-labelled antamanide

The effect of the crystal lattice on the side-chain conformation andside-chain dynamics in peptides is investigated by solid-state NMR, using thecyclic decapeptide antamanide as an example. The study takes advantage of the¹³C assignment of the backbone and side chains based on theresolution-enhanced 2D spin-diffusion spectra by heteronuclear and homonucleardecoupling. The spectra even allow for a stereospecific assignment of the-carbons of the valine residue. It is found that the valine side chaincoexists in two static rotamer conformations which have not been observed byX-ray crystallography. In addition, remarkable effects of the crystal packingon the methyl-group rotation frequency are found from ¹³Crelaxation measurements.     

Coos,booms, and hoots: The evolution of closed‐mouth vocal behavior in birds

Most birds vocalize with an open beak, but vocalization with a closed beak into an inflating cavity occurs in territorial or courtship displays in disparate species throughout birds. Closed‐mouth vocalizations generate resonance conditions that favor low‐frequency sounds. By contrast, open‐mouth vocalizations cover a wider frequency range. Here we describe closed‐mouth vocalizations of birds from functional and morphological perspectives and assess the distribution of closed‐mouth vocalizations in birds and related outgroups. Ancestral‐state optimizations of body size and vocal behavior indicate that closed‐mouth vocalizations are unlikely to be ancestral in birds and have evolved independently at least 16 times within Aves, predominantly in large‐bodied lineages. Closed‐mouth vocalizations are rare in the small‐bodied passerines. In light of these results and body size trends in nonavian dinosaurs, we suggest that the capacity for closed‐mouth vocalization was present in at least some extinct nonavian dinosaurs. As in birds, this behavior may have been limited to sexually selected vocal displays, and hence would have co‐occurred with open‐mouthed vocalizations.