Chemical rescue of a site-modified ligand to a [4Fe-4S] cluster in PsaC, a bacterial-like dicluster ferredoxin bound to Photosystem I

Chemical rescue of site-modified amino acids using externally supplied organic molecules represents a powerful method to investigate structure-function relationships in proteins. Here we provide definitive evidence that aryl and alkyl thiolates, reagents typically used for in vitro iron-sulfur cluster reconstitutions, serve as rescue ligands to a site-specifically modified [4Fe-4S]^1+,2+ cluster in PsaC, a bacterial dicluster ferredoxin-like subunit of Photosystem I. PsaC binds two low-potential [4Fe-4S]^1+,2+ clusters termed F_A and F_B. In the C13G/C33S variant of PsaC, glycine has replaced cysteine at position 13 creating a protein that is missing one of the ligating amino acids to iron-sulfur cluster F_B. Using a variety of analytical techniques, including non-heme iron and acid-labile sulfur assays, and EPR, resonance Raman, and Mössbauer spectroscopies, we showed that the C13G/C33S variant of PsaC binds two [4Fe-4S]^1+,2+ clusters, despite the absence of one of the biological ligands. ¹⁹F NMR spectroscopy indicated that the external thiolate replaces cysteine 13 as a substitute ligand to the F_B cluster. The finding that site-modified [4Fe-4S]^1+,2+ clusters can be chemically rescued with external thiolates opens new opportunities for modulating their properties in proteins. In particular, it provides a mechanism to attach an additional electron transfer cofactor to the protein via a bound, external ligand.     

Phosphoproteome Profiling of the Receptor Tyrosine Kinase MuSK Identifies Tyrosine Phosphorylation of Rab GTPases

Muscle-specific receptor tyrosine kinase (MuSK) agonist antibodies were developed 2 decades ago to explore the benefits of receptor activation at the neuromuscular junction. Unlike agrin, the endogenous agonist of MuSK, agonist antibodies function independently of its coreceptor low-density lipoprotein receptor–related protein 4 to delay the onset of muscle denervation in mouse models of ALS. Here, we performed dose–response and time-course experiments on myotubes to systematically compare site-specific phosphorylation downstream of each agonist. Remarkably, both agonists elicited similar intracellular responses at known and newly identified MuSK signaling components. Among these was inducible tyrosine phosphorylation of multiple Rab GTPases that was blocked by MuSK inhibition. Importantly, mutation of this site in Rab10 disrupts association with its effector proteins, molecule interacting with CasL 1/3. Together, these data provide in-depth characterization of MuSK signaling, describe two novel MuSK inhibitors, and expose phosphorylation of Rab GTPases downstream of receptor tyrosine kinase activation in myotubes.     

Classification of β-hairpin repeat proteins

In recent years, a number of new protein structures that possess tandem repeats have emerged. Many of these proteins are comprised of tandem arrays of β-hairpins. Today, the amount and variety of the data on these β-hairpin repeat (BHR) structures have reached a level that requires detailed analysis and further classification. In this paper, we classified the BHR proteins, compared structures, sequences of repeat motifs, functions and distribution across the major taxonomic kingdoms of life and within organisms. As a result, we identified six different BHR folds in tandem repeat proteins of Class III (elongated structures) and one BHR fold (up-and-down β-barrel) in Class IV (“closed” structures). Our survey reveals the high incidence of the BHR proteins among bacteria and viruses and their possible relationship to the structures of amyloid fibrils. It indicates that BHR folds will be an attractive target for future structural studies, especially in the context of age-related amyloidosis and emerging infectious diseases. This work allowed us to update the RepeatsDB database, which contains annotated tandem repeat protein structures and to construct sequence profiles based on BHR structural alignments.     

似鳡属的系统地位及种的分化(鲤形目:鲤科)

似鳡属(Luciocyprinus Vaillant)是鲤科鱼类中属于鲃系(陈湘粦等,1984)的一个属,下咽齿3行,臀鳍无硬刺,具分枝鳍条5根。过去只记载一个种,外形很像鳡鱼,是由于摄食习性相同而导致的趋同现象(林人端,1981),故称似鳡,在广西俗称墨线鳡,云南俗称杆条鱼。分布于西江水系,包括云南中南部的湖泊,与鳡鱼分布于同一水系,但各占一定的水域,互不重叠。 关于似鳡的拉丁名称,惯用Fustis vivus Lin,是林书颜1932年定的新属新种,     

Oxidized albumin. The long way of a protein of uncertain function

Background

Proteins are extremely reactive to oxidants and should represent a potential target of instable reactive oxygen. This may represent a problem for plasma proteins since they may be directly modified in vivo in a compartment where antioxidant enzymatic systems are scarcely represented. On the other hand, it is possible that some plasma components have evolved over time to guarantee protection, in which case they can be considered as anti-oxidants.

Scope of review

To present and discuss main studies which addressed the role of albumin in plasma antioxidant activity mainly utilizing in vitro models of oxidation. To present some advances on structural features of oxidized albumin deriving from studies carried out on in vitro models as well as albumin purified in vivo from patients affected by clinical conditions characterized by oxidative stress.

Major conclusions

There are different interaction with HOCl and chloramines. In the former case, HOCl produces an extensive alteration of ²³⁸Trp and ¹⁶²Tyr, ⁴²⁵Tyr, ⁴⁷Tyr, while thiol groups are only partially involved. Chloramines are extremely reactive with the unique free SH group of albumin (³⁴Cys) with the formation of sulfenic and sulfinic acid as intermediates and sulfonic acid as end-product. Oxidized albumin has a modified electrical charge for the addition of an acidic residue and presents α-helix and random coil reorganization with subtle changes in domain orientation.

General significance

Albumin, is the major antioxidants in plasma with a concentration (0.8 mM) higher than other antioxidants by an exponential factor. Functional and protective roles in the presence of oxidative stress must be defined. This article is part of a Special Issue entitled Serum Albumin.     

Effects of the cardiomyopathy-causing E244D mutation of troponin T on the structures of cardiac thin filaments studied by small-angle X-ray scattering

Small-angle X-ray scattering experiments were carried out to investigate the structural changes of cardiac thin filaments induced by the cardiomyopathy-causing E244D mutation in troponin T (TnT). We examined native thin filaments (NTF) from a bovine heart, reconstituted thin filaments containing human cardiac wild-type Tn (WTF), and filaments containing the E244D mutant of Tn (DTF), in the absence and presence of Ca²⁺. Analysis by model calculation showed that upon Ca²⁺-activation, tropomyosin (Tm) and Tn in the WTF and NTF moved together in a direction to expose myosin-binding sites on actin. On the other hand, Tm and Tn of the DTF moved in the opposite directions to each other upon Ca²⁺-activation. These movements caused Tm to expose more myosin-binding sites on actin than the WTF, suggesting that the affinity of myosin for actin is higher for the DTF. Thus, the mutation-induced structural changes in thin filaments would increase the number of myosin molecules bound to actin compared with the WTF, resulting in the force enhancement observed for the E244D mutation.     

Simple sequence repeat genetic linkage maps of A-genome diploid cotton (Gossypium arboreum)

This study introduces the construction of the first intraspacific genetic linkage map of the A-genome diploid cotton with newly developed simple sequence repeat (SSR) markers using 189 F2 plants derived from the cross of two Asiatic parents were detected using 6 092 pairs of SSR primers. Two-hundred and sixty-eight pairs of SSR pdmers with better polymorphisms were picked out to analyze the F2 population. In total, 320 polymorphic bands were generated and used to construct a linkage map with JoinMap3.0. Two-hundred and sixty-seven loci, Including three phenotypic traits were mapped at a logarithms of odds ratio (LOD) ≥ 3.0 on 13 linkage groups. The total length of the map was 2 508.71 cM, and the average distance between adjacent markers was 9.40 cM. Chromosome assignments were according to the association of linkages with our backbone tetraploid specific map using the 89 similar SSR loci. Comparisons among the 13 suites of orthologous linkage groups revealed that the A-genome chromosomes are largely collinear with the At and Dt sub-genome chromosomes. Chromosomes associated with inversions suggested that allopolyploidization was accompanied by homologous chromosomal rearrangement. The inter-chromosomal duplicated loci supply molecular evidence that the A-genome diploid Asiatic cotton is paleopolyploid.     

PONDR-FIT: A meta-predictor of intrinsically disordered amino acids

Protein intrinsic disorder is becoming increasingly recognized in proteomics research. While lacking structure, many regions of disorder have been associated with biological function. There are many different experimental methods for characterizing intrinsically disordered proteins and regions; nevertheless, the prediction of intrinsic disorder from amino acid sequence remains a useful strategy especially for many large-scale proteomic investigations. Here we introduced a consensus artificial neural network (ANN) prediction method, which was developed by combining the outputs of several individual disorder predictors. By eight-fold cross-validation, this meta-predictor, called PONDR-FIT, was found to improve the prediction accuracy over a range of 3 to 20% with an average of 11% compared to the single predictors, depending on the datasets being used. Analysis of the errors shows that the worst accuracy still occurs for short disordered regions with less than ten residues, as well as for the residues close to order/disorder boundaries. Increased understanding of the underlying mechanism by which such meta-predictors give improved predictions will likely promote the further development of protein disorder predictors. Access to PONDR-FIT is available at www.disprot.org.