Isoprenoids constitute one of the most diverse classes of natural products. As a compound class, they are essential to basic metabolic processes including cell-wall biosynthesis, post-translational ...protein modifications, and signaling. In addition, isoprenoid secondary metabolites are highly valuable natural products with a wide range of biotechnological applications. The biosynthesis of their two universal building blocks, isopentenyl diphosphate and dimethylallyl diphosphate, was thought to proceed exclusively by way of mevalonate as a key intermediate until a novel pathway involving methylerithritol phosphate (MEP) was discovered in the early 1990s. In this review, we describe the seven enzymes of the MEP pathway, along with their discoveries, three-dimensional structures, and mechanisms. The latter include examples of remarkable enzyme catalysis including an unusual cytidilation reaction and the use of iron–sulfur cluster cofactors in reductive ring opening and hydroxy-group elimination. Furthermore, isoprenoid biosynthesis shows a characteristic species distribution. A brief overview highlights the MEP pathway’s potential as a selective drug target, which is absent in humans but essential to the survival of many important bacterial and apicomplexan pathogens.
Proteolytic degradation is an essential cellular process which is primarily carried out by the 20S proteasome core particle (CP), a protease of 720 kDa and 28 individual subunits. As a result of its ...central functional role, the proteasome represents an attractive drug target that has been extensively investigated during the last decade and validated by the approval of bortezomib by the US Food and Drug Administration (FDA). Currently, several optimized second‐generation proteasome inhibitors are being explored as anticancer drugs in clinical trials, and most of them target both constitutive proteasomes (cCPs) and immunoproteasomes (iCPs). However, selective inhibition of the iCPs, a distinct class of proteasomes predominantly expressed in immune cells, appears to be a promising therapeutic rationale for the treatment of autoimmune disorders. Although a few selective agents have already been identified, the recently determined crystal structure of the iCP will further promote the development and optimization of iCP‐selective compounds.
Under arrest: The first immunoproteasome‐specific inhibitors were recently developed. The most potent one shows promise in the treatment of autoimmune diseases, thus opening the door for new clinical applications. Structure–affinity studies with the available immuno‐ and constitutive proteasome crystal structures will facilitate future drug development efforts.
The heart of the ubiquitin-mediated degradation pathway, the 26S proteasome, endoproteolytically cleaves most intracellular proteins, thereby maintaining biological homeostasis and regulating many ...crucial processes in the cell. This hydrolyzing machine comprises more than 30 different subunits, which perform different functions including the recognition, unfolding, translocating and cleavage of protein substrates. Thus, careful assemblage and regulation of the 26S proteasome is essential to ensure correct positioning and function of each subunit, thereby preserving the delicate cellular balance between protein synthesis and degradation. Here, we review the most current research on the 26S proteasome assembly pathway, and describe the mechanism used by the cell to manage the complex structure and functions of the proteasome.
This article discusses the structure and function of the proteasome and its inhibitors. The proteasome is playing a significant role in the turnover of cellular proteins.
•We investigated the intra-annual runoff variations and periods in an endorheic lake basin.•The runoff quantities were investigated by using structuring models.•Both climate variability and ...anthropogenic activities had effects on the total river runoff.•The climate variability was a leading factor driving runoff evolution in the lake Issyk-Kul basin.
An attribute study of the runoff changes due to climate change and anthropogenic activities is of great value to determining the best response to runoff changes. This study investigates the evolution of the intra-annual runoff and evaluates the effects of changes in climate and anthropogenic activities on runoff variations in the Lake Issyk-Kul Basin (LIKB). The monotonic trend test and wavelet transform analysis were used to characterize the trends and periods of the hydro-meteorological variables in the LIKB. In addition, the periodicity-trend superposition model and double-mass curves were used to assess the contributions of climate change and anthropogenic activities to runoff variations. The analyses were conducted using regional climate data combined with runoff data derived from 13 different river stations in the LIKB for the period between 1943 and 2012. The results indicate the following. (1) The monthly runoff trends in the three groups of tributaries (northern, southeastern, and southern tributaries) exhibit distinct differences, and the percentage of the data sets with no pattern gradually increased (from 26% to 38%) from north to south. There is a positive trend, with a similar decrease (from 50% to 24%) from north to south. (2) The air temperature, precipitation, and runoff in the LIKB increased throughout the low and high flow periods. The statistical index of the runoff concentration degree (RCD) decreased at most stations (Chong-Koi-Suu, Chong-Ak-Suu, Chong-Urykty, Pzhergalan, Ak-Sai, Tossor, Ton, Chong-Kyzyl-Suu, Tamga, and Karakol), except for the Cholpon-Ata, Ak-Suu, and Dzhuuku stations. (3) In addition, the periodic characteristics of the RCD show that the RCD has a 12-year significant period over the low flow period, which is similar to that for precipitation but varies from that for air temperature. (4) Climate variability and anthropogenic activities are two different contributors to runoff reduction in the LIKB. The respective contributions of anthropogenic activities to the runoff determined using the two methods were 35.3% and 18%, whereas the climate change contributions were 64.7% and 82%, respectively. Climate change was the main reason for the runoff variations. These results provide a basis for the scientific management and sustainable allocation of water resources in the LIKB.
The heptameric proteasome activator (PA) 28αβ is known to modulate class I antigen processing by docking onto 20S proteasome core particles (CPs). The exact stoichiometry and arrangement of its α and ...β subunits, however, is still controversial. Here we analyzed murine PA28 complexes regarding structure and assembly. Strikingly, PA28α, PA28β, and PA28αβ preparations form heptamers, but solely PA28α and PA28αβ associate with CPs. Co-expression of α and β yields one unique PA28αβ species with an unchangeable subunit composition. Structural data on PA28α, PA28β, and PA28αβ up to 2.9 Å resolution reveal a PA28α4β3 complex with an alternating subunit arrangement and a single α-α interface. Differential scanning fluorimetry experiments and activity assays classify PA28α4β3 as most stable and most active, indicating that this assembly might represent the physiologically relevant species. Together, our data resolve subunit composition and arrangement of PA28αβ and clarify how an asymmetric heptamer can be assembled from two highly homologous subunits.
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•Crystal structures of mouse PA28α7, PA28β7, and PA28α4β3 were determined•Despite forming a heptamer PA28β fails to bind to proteasomes•The alternating arrangement of four α and three β subunits in PA28αβ is unchangeable•β-β and α-α contacts are less stable than α-β interfaces
PA28 complexes activate 20S proteasomes and modulate their product cleavage patterns. Huber and Groll crystallized and biochemically analyzed mouse PA28α, PA28β, and PA28αβ preparations. Being assembled of four α and three β subunits, PA28α4β3 is more stable and more active than PA28α7 and PA28β7.
Constitutive proteasomes and immunoproteasomes shape the peptide repertoire presented by major histocompatibility complex class I (MHC-I) molecules by harboring different sets of catalytically active ...subunits. Here, we present the crystal structures of constitutive proteasomes and immunoproteasomes from mouse in the presence and absence of the epoxyketone inhibitor PR-957 (ONX 0914) at 2.9 Å resolution. Based on our X-ray data, we propose a unique catalytic feature for the immunoproteasome subunit β5i/LMP7. Comparison of ligand-free and ligand-bound proteasomes reveals conformational changes in the S1 pocket of β5c/X but not β5i, thereby explaining the selectivity of PR-957 for β5i. Time-resolved structures of yeast proteasome:PR-957 complexes indicate that ligand docking to the active site occurs only via the reactive head group and the P1 side chain. Together, our results support structure-guided design of inhibitory lead structures selective for immunoproteasomes that are linked to cytokine production and diseases like cancer and autoimmune disorders.
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► First ligand structures of mouse constitutive proteasomes and immunoproteasomes are determined ► A unique hydrophilic architecture surrounds the β5i-oxyanion hole ► Structures explain the enhanced generation of MHC-I antigens by immunoproteasomes ► The mode of action and selectivity of the β5i-specific compound PR-957 is revealed
Structural insights into the molecular differences between the constitutive proteasome and the immunoproteasome explain why the immunoproteasome is more competent in producing MHC-I ligands and why it is preferentially targeted by the inhibitor PR-957.
The analysis of various characteristics and trends of precipitation is an essential task to improve the utilization of water resources. Lake Issyk-Kul basin is an upper alpine catchment, which is ...more susceptible to the effects of climate variability, and identifying rainfall variations has vital importance for water resource planning and management in the lake basin. The well-known approaches linear regression, Şen’s slope, Spearman’s rho, and Mann-Kendall trend tests are applied frequently to try to identify trend variations, especially in rainfall, in most literature around the world. Recently, a newly developed method of Şen-innovative trend analysis (ITA) provides some advantages of visual-graphical illustrations and the identification of trends, which is one of the main focuses in this article. This study obtained the monthly precipitation data (between 1951 and 2012) from three meteorological stations (Balykchy, Cholpon-Ata, and Kyzyl-Suu) surrounding the Lake Issyk-Kul, and investigated the trends of precipitation variability by applying the ITA method. For comparison purposes, the traditional Mann–Kendall trend test also used the same time series. The main results of this study include the following. (1) According to the Mann-Kendall trend test, the precipitation of all months at the Balykchy station showed a positive trend (except in January (Zc = −0.784) and July (Zc = 0.079)). At the Cholpon-Ata and Kyzyl-Suu stations, monthly precipitation (with the same month of multiple years averaged) indicated a decreasing trend in January, June, August, and November. At the monthly scale, significant increasing trends (Zc > Z0.10 = 1.645) were detected in February and October for three stations. (2) The ITA method indicated that the rising trends were seen in 16 out of 36 months at the three stations, while six months showed decreasing patterns for “high” monthly precipitation. According to the “low” monthly precipitations, 14 months had an increasing trend, and four months showed a decreasing trend. Through the application of the ITA method (January, March, and August at Balykchy; December at Cholpon-Ata; and July and December at Kyzyl-Suu), there were some significant increasing trends, but the Mann-Kendall test found no significant trends. The significant trend occupies 19.4% in the Mann-Kendall test and 36.1% in the ITA method, which indicates that the ITA method displays more positive significant trends than Mann–Kendall Zc. (3) Compared with the classical Mann-Kendall trend results, the ITA method has some advantages. This approach allows more detailed interpretations about trend detection, which has benefits for identifying hidden variation trends of precipitation and the graphical illustration of the trend variability of extreme events, such as “high” and “low” values of monthly precipitation. In contrast, these cannot be discovered by applying traditional methods.
Covalent DNA-protein crosslinks (DPCs) are toxic DNA lesions that interfere with essential chromatin transactions, such as replication and transcription. Little was known about DPC-specific repair ...mechanisms until the recent identification of a DPC-processing protease in yeast. The existence of a DPC protease in higher eukaryotes is inferred from data in Xenopus laevis egg extracts, but its identity remains elusive. Here we identify the metalloprotease SPRTN as the DPC protease acting in metazoans. Loss of SPRTN results in failure to repair DPCs and hypersensitivity to DPC-inducing agents. SPRTN accomplishes DPC processing through a unique DNA-induced protease activity, which is controlled by several sophisticated regulatory mechanisms. Cellular, biochemical, and structural studies define a DNA switch triggering its protease activity, a ubiquitin switch controlling SPRTN chromatin accessibility, and regulatory autocatalytic cleavage. Our data also provide a molecular explanation on how SPRTN deficiency causes the premature aging and cancer predisposition disorder Ruijs-Aalfs syndrome.
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•The SPRTN metalloprotease repairs DNA-protein crosslinks•A DNA switch controls SPRTN’s protease activity•A ubiquitin switch controls chromatin access of SPRTN•Structural insights reveal unique features of the SPRTN/Wss1 protease family
Stingele et al. discover the SPRTN metalloprotease to be crucial for DNA-protein crosslink repair in higher eukaryotes. In addition, several regulatory principles constraining SPRTN’s potentially toxic activity are described: a ubiquitin switch controlling chromatin access, a DNA switch triggering protease activity, and a negative feedback loop based on autocatalytic cleavage.
Biogenesis of the 20S proteasome is tightly regulated. The N-terminal propeptides protecting the active-site threonines are autocatalytically released only on completion of assembly. However, the ...trigger for the self-activation and the reason for the strict conservation of threonine as the active site nucleophile remain enigmatic. Here we use mutagenesis, X-ray crystallography and biochemical assays to suggest that Lys33 initiates nucleophilic attack of the propeptide by deprotonating the Thr1 hydroxyl group and that both residues together with Asp17 are part of a catalytic triad. Substitution of Thr1 by Cys disrupts the interaction with Lys33 and inactivates the proteasome. Although a Thr1Ser mutant is active, it is less efficient compared with wild type because of the unfavourable orientation of Ser1 towards incoming substrates. This work provides insights into the basic mechanism of proteolysis and propeptide autolysis, as well as the evolutionary pressures that drove the proteasome to become a threonine protease.
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