Creating a community resource for protein science

Helen Berman is the recipient of the Protein Society 2012 Carl Branden Award In addition to being one of the early pioneers in protein crystallography, Carl Brändén made significant contributions to science education with his elegant and beautifully illustrated book Introduction to Protein Structure (Brändén and Tooze, New York: Garland, 1991). It is truly an honor to receive this award in their names. This award and the 40th anniversary of the Protein Data Bank (PDB; Berman et al., Structure 2012;20:391–396) have given me an opportunity to reflect on the various components that have contributed to building a resource for protein science and to try to quantify the impact of having PDB data openly available.     

Induction of the unfolded protein response in familial amyotrophic lateral sclerosis and association of protein-disulfide isomerase with superoxide dismutase 1

Mutations in Cu/Zn superoxide dismutase (SOD1) are linked to motor neuron death in familial amyotrophic lateral sclerosis (ALS) by an unclear mechanism, although misfolded SOD1 aggregates are commonly associated with disease. Proteomic analysis of the transgenic SOD1(G93A) ALS rat model revealed significant up-regulation of endoplasmic reticulum (ER)-resident protein-disulfide isomerase (PDI) family members in lumbar spinal cords. Expression of SOD1 mutants (mSOD1) led to an up-regulation of PDI in motor neuron-like NSC-34 cells but not other cell lines. Inhibition of PDI using bacitracin increased aggregate production, even in wild type SOD1 transfectants that do not readily form inclusions, suggesting PDI may protect SOD1 from aggregation. Moreover, PDI co-localized with intracellular aggregates of mSOD1 and bound to both wild type and mSOD1. SOD1 was also found in the microsomal fraction of cells despite being a predominantly cytosolic enzyme, confirming ER-Golgi-dependent secretion. In SOD1(G93A) mice, a significant up-regulation of unfolded protein response entities was also observed during disease, including caspase-12, -9, and -3 cleavage. Our findings therefore implicate unfolded protein response and ER stress-induced apoptosis in the patho-physiology of familial ALS. The possibility that PDI may be a therapeutic target to prevent SOD1 aggregation is also raised by this study.     

Isolation and properties of the soluble c-type cytochromes of the dinoflagellate Peridinium cinctum

Four soluble cytochromes of the c type were isolated from the freshwater dinoflagellate Peridinium cinctum collected from Lake Kinneret, Israel. Cytochrome c with alpha-band maximum at 550 nm in the reduced state had a molecular mass of 10,200 Da, pI 7.4, and Em of 278 m V. This cytochrome was active in the respiratory chain of beef heart Keilin-Hartree particles. Cytochrome c-553 had a molecular mass of 13,200 Da, pI 4.9, and Em of 384 m V, and was active in light induced electron transport of Euglena gracilis chloroplast fragments. Cytochrome c-554 had a molecular mass of 13,500 Da, pI 4.4, and Em of 326 m V. This cytochrome was inactive in light induced electron transport but competed with cytochrome c-552 of Euglena in the assay. The acidic cytochrome c-557 was present in very small quantities. The properties of the soluble c-type cytochromes of P. cinctum are compatible with the classification of dinoflagellates as primitive eucaryotes.     

��-arrestin Kurtz inhibits MAPK and Toll signalling in Drosophila development

β‐Arrestins have been implicated in the regulation of multiple signalling pathways. However, their role in organism development is not well understood. In this study, we report a new in vivo function of the Drosophila β‐arrestin Kurtz (Krz) in the regulation of two distinct developmental signalling modules: MAPK ERK and NF‐κB, which transmit signals from the activated receptor tyrosine kinases (RTKs) and the Toll receptor, respectively. Analysis of the expression of effectors and target genes of Toll and the RTK Torso in krz maternal mutants reveals that Krz limits the activity of both pathways in the early embryo. Protein interaction studies suggest a previously uncharacterized mechanism for ERK inhibition: Krz can directly bind and sequester an inactive form of ERK, thus preventing its activation by the upstream kinase, MEK. A simultaneous dysregulation of different signalling systems in krz mutants results in an abnormal patterning of the embryo and severe developmental defects. Our findings uncover a new in vivo function of β‐arrestins and present a new mechanism of ERK inhibition by the Drosophila β‐arrestin Krz.     

Engineering glycoproteins for use as pharmaceuticals

The fact that glycosylation is not a significant process in prokaryotes means that many of the proteins produced by genetically engineered bacteria are not identical to their eukaryotic counterparts. Although glycosylation affects the physical, chemical and biological nature of proteins, its pharmacological value in potential protein pharmaceuticals is not easy to predict. However, the development of mammalian cell culture methods for expressing recombinant DNA-derived glycoproteins will permit further studies in the field.     

Correction to: Managing biological invasions: the cost of inaction

Biological Invasions -     

The ontogeny and population variability of human hepatic dihydronicotinamide riboside:quinone oxidoreductase (NQO2)

Dihydronicotinamide riboside:quinone oxidoreductase (NQO2) is an enzyme that performs reduction reactions involved in antioxidant defense. We hypothesized that NQO2 hepatic drug clearance would develop in children over time, similar to NQO1. Using human liver cytosol (n = 117), the effects of age, sex, ethnicity, and weight on NQO2 expression and activity were probed. No significant correlations were observed. Biochemical activity of NQO2 was as high at birth as in adults (0.23 ± 0.04 nmol/min/mg protein, mean ± SEM, range 0–1.83). In contrast, modeled hepatic clearance through the NQO2 pathway was up to 10% of adult levels at birth, reaching predicted adult levels (0.3 ± 0.03 L/h) at 14 years of age. Comparisons between NQO1 and NQO2 in the same livers showed that neither protein (P = 0.32) nor activity (P = 0.23) correlated, confirming both orthologs are independently regulated. Because hepatic clearance through NQO2 does not mature until teenage years, compounds detoxified by this enzyme may be more deleterious in children.     

Metabolic control analysis is helpful for informed genetic manipulation of oilseed rape (Brassica napus) to increase seed oil content

Top-down control analysis (TDCA) is a useful tool for quantifying constraints on metabolic pathways that might be overcome by biotechnological approaches. Previous studies on lipid accumulation in oilseed rape have suggested that diacylglycerol acyltransferase (DGAT), which catalyses the final step in seed oil biosynthesis, might be an effective target for enhancing seed oil content. Here, increased seed oil content, increased DGAT activity, and reduced substrate:product ratio are demonstrated, as well as reduced flux control by complex lipid assembly, as determined by TDCA in Brassica napus (canola) lines which overexpress the gene encoding type-1 DGAT. Lines overexpressing DGAT1 also exhibited considerably enhanced seed oil content under drought conditions. These results support the use of TDCA in guiding the rational selection of molecular targets for oilseed modification. The most effective lines had a seed oil increase of 14%. Moreover, overexpression of DGAT1 under drought conditions reduced this environmental penalty on seed oil content.     

An examination of targeted gene neighborhoods in strawberry

Background  

Strawberry (Fragaria spp.) is the familiar name of a group of economically important crop plants and wild relatives that also represent an emerging system for the study of gene and genome evolution. Its small stature, rapid seed-to-seed cycle, transformability and miniscule basic genome make strawberry an attractive system to study processes related to plant physiology, development and crop production; yet it lacks substantial genomics-level resources. This report addresses this deficiency by characterizing 0.71 Mbp of gene space from a diploid species (F. vesca). The twenty large genomic tracks (30-52 kb) captured as fosmid inserts comprise gene regions with roles in flowering, disease resistance, and metabolism.