Darwin's apes and “savages”

Since his visit to Tierra del Fuego in the 1830s, Darwin had been fascinated by the “savages” that succeeded in surviving on such a “broken beach”, and because they were certainly similar in behaviour to our ancestors. However, he was also fascinated by baboons’ behaviour, according to Brehm's accounts: hamadryas baboons showed a strong altruism to the point of risking their own lives in order to save their infants from attack by dogs. In 1871, he mentions he would rather have descended from brave baboons than from “savages”, considered egoistic. We study the two sources of these ideas and try to show how Darwin's comparative reflections on apes and “savages” made him the first evolutionist anthropologist.     

Structure of Internalin InlK from the Human Pathogen Listeria monocytogenes

Listeria monocytogenes is a human pathogen that employs a wide variety of virulence factors in order to adhere to, invade, and replicate within target cells. Internalins play key roles in processes ranging from adhesion to receptor recognition and are thus essential for infection. Recently, InlK, a surface-associated internalin, was shown to be involved in Listeria's ability to escape from autophagy by recruitment of the major vault protein (MVP) to the bacterial surface. Here, we report the structure of InlK, which harbors four domains arranged in the shape of a “bent arm”. The structure supports a role for the “elbow” of InlK in partner recognition, as well as of two Ig-like pedestals intercalated by hinge regions in the projection of InlK away from the surface of the bacterium. The unusual fold and flexibility of InlK could be essential for MVP binding and concealment from recognition by molecules involved in the autophagic process.     

The state transition mechanism—simply depending on light-on and -off in Spirulina platensis

The state transition in cyanobacteria is a long-discussed topic of how the photosynthetic machine regulates the excitation energy distribution in balance between the two photosystems. In the current work, whether the state transition is realized by “mobile phycobilisome (PBS)” or “energy spillover” has been clearly answered by monitoring the spectral responses of the intact cells of the cyanobacterium Spirulina platensis. Firstly, light-induced state transition depends completely on a movement of PBSs toward PSI or PSII while the redox-induced one on not only the “mobile PBS” but also an “energy spillover”. Secondly, the “energy spillover” is triggered by dissociation of PSI trimers into the monomers which specially occurs under a case from light to dark, while the PSI monomers will re-aggregate into the trimers under a case from dark to light, i.e., the PSI oligomerization is reversibly regulated by light switch on and off. Thirdly, PSI oligomerization is regulated by the local H⁺ concentration on the cytosol side of the thylakoid membranes, which in turn is regulated by light switch on and off. Fourthly, PSI oligomerization change is the only mechanism for the “energy spillover”. Thus, it can be concluded that the “mobile PBS” is a common rule for light-induced state transition while the “energy spillover” is only a special case when dark condition is involved.     

Characterization and polymorphism of Keratin Associated Protein 1.4 gene in goats

Keratin-associated proteins (KAPs) are among the main structural components of the animal fibers and form semi-rigid matrix wherein the keratin intermediate filaments (KIFs) are embedded. Variation in the KAP genes has been reported to affect the structure of KAPs and hence fiber characteristics. As no information is available on this gene in Capra hircus therefore, present work was undertaken to characterize and explore the different polymorphic variants of KAP1.4 gene at DNA level in different breeds/genetic groups of goats of Kashmir. Cashmere (Changthangi, 30 animals) and non-Cashmere (Bakerwal and Kargil goats, 20 animals each) goats formed the experimental animals for the study. Single strand conformation polymorphism technique was employed for exploring variability at gene level. On exploring the size variability in KAP1.4 gene between Ovine and Caprine, it was concluded that sheep KAP1.4 gene has a deletion of 30 nucleotides. In comparison to published nucleotide sequences of sheep, goat sequences explored are differing at positions 174, 462 and 568 and at these positions “G”, “T” and “T” nucleotides are present in sheep, but are replaced by “A”, “C” and “C” respectively, in goats. By SSC studies, two genotypes were observed in each genetic group and in Bakerwal goats the genotypes were designated as A1A1 (0.40) and A1A2 (0.60) and were formed by two alleles A1 (0.70) andA2 (0.30). The different SSC patterns observed in Kargil goats were designated as B1B1 (0.35) and B1B2 (0.65) genotypes with frequencies of B1 and B2 alleles as 0.675 and 0.325, respectively. Similarly, two genotypes C1C1 (0.60) and C1C2 (0.40) were observed in Changthangi goats and the frequencies of C1 and C2 alleles were 0.80 and 0.20, respectively. These alleles were later confirmed by sequencing. The sequences of these alleles are available in NCBI under Acc. No's. JN012101.1, JN012102.1, JN000317.1, JN000318.1, JQ436929 and JQ627657. It was concluded that all the alleles observed in a breed were unique to the breed. The designated A1 and A2 alleles of Bakerwal goats differ from each other at positions 245 and the nucleotides observed were “C” or “A” and at position 605 of the nucleotide sequence “T” or “C”, were observed. The designated B1 and B2 alleles of Kargil goats differed from each other at positions 224, 374, 375 and 521. The nucleotides observed in two SSC pattern were C→G, A→G, G→A and T→C, respectively. The designated C1 and C2 alleles of Changthangi goats differed from each other at one position 440 with the change of “A”→“C”.     

Darwin teleologist? Design in the Orchids

Focusing on the Orchids, this article aims at disentangling the concepts of teleology, design and natural theology. It refers to several contemporary critics of Darwin (Kölliker, Argyll, Royer, Candolle, Delpino) to challenge Huxley's interpretation that Darwin's system was “a deathblow” to teleology. The Orchids seem rather to be a “flank-movement” (Gray): it departs from the Romantic theories of transmutation and the “imaginary examples” of the Origin; it focuses on empirical data and on teleological structures. Although Darwin refers to natural selection, his readers mock him for his fascination for delicate morphological contrivances and co-adaptations – a sign that he was inescapably lured to finality. Some even suggested that his system was a “theodicy”. In the history of Darwinism, the Orchids reveal “another” quite unexpected and heterodox Darwin: freed from the hypothetical fancies of the Origin, and even suggesting a new kind of physico-theology.     

Mechanisms of cohesin-mediated gene regulation and lessons learned from cohesinopathies

Cohesins are conserved and essential Structural Maintenance of Chromosomes (SMC) protein-containing complexes that physically interact with chromatin and modulate higher-order chromatin organization. Cohesins mediate sister chromatid cohesion and cellular long-distance chromatin interactions affecting genome maintenance and gene expression. Discoveries of mutations in cohesin's subunits and its regulator proteins in human developmental disorders, so-called “cohesinopathies,” reveal crucial roles for cohesins in development and cellular growth and differentiation. In this review, we discuss the latest findings concerning cohesin's functions in higher-order chromatin architecture organization and gene regulation and new insight gained from studies of cohesinopathies. This article is part of a Special Issue entitled: Chromatin and epigenetic regulation of animal development.     

Analysis of protein aggregation kinetics using short amino acid peptide tags

Understanding protein solubility, and consequently aggregation, is an important issue both from an academic and a biotechnological application viewpoints. Here we report the effects of 10 representative amino acids on the aggregation kinetics of proteins. The effects were determined by measuring the solubility of a simplified bovine pancreatic trypsin inhibitor (BPTI) variant, to which short artificial tags containing the amino acid of interest were added at its C-terminus. We determined the solubility of the tagged variants as a function of equilibration time (20 min to 48 h) and total protein concentration ranging from 0.10 mg/ml to 25.0 mg/ml. We observed, as anticipated, that proteins precipitated when the total protein concentration exceeded a critical value. However, when the total protein concentration was further increased, the apparent solubility reached a concentration above the critical value, and slowly decreased to a value under the critical concentration upon increasing the equilibration period. We rationalized these observations by identifying three different solubility values, the “transient solubility (TS)”, the “aggregation initiation concentration (AIC)” and the “long-term solubility (LS)”. AIC and LS are parameters determined essentially by the amino acid types composing the tags and could be considered as an amino acid's intrinsic property. On the other hand, TS is an apparent solubility that is measured after some (20 min in our case) equilibration time and is often considered as the “solubility” of the protein. Similar aggregation kinetic patterns were observed with natural proteins, indicating the generality of the observations made using our model protein.     

Amino-terminal domain interactions of lambda integrase on arm-type DNA

In contrast to the other tyrosine recombinase family members, integrase protein (Int) of bacteriophage λ has an additional amino-terminal domain that binds to “arm-type” DNA sequences distant from those involved in strand exchange. The homomeric interaction between neighboring amino-terminal domains of Int is contributed by R30-D71 salt-bridge in a non-equivalent manner on Holliday-junction intermediates. In this report, R30 and D71 residues were investigated in regard to Int’s cooperative binding to “arm-type” DNA and the attenuating function of “arm-type” DNA. The results suggest the electrostatic interaction between residues 30 and 71 is dependent on “arm-type” DNA and contributes the “selective” inhibition of catalytic activity of λ Int by “arm-type” DNA.     

Atypical yeasts identified as Saccharomyces cerevisiae by MALDI-TOF MS and gene sequencing are the main responsible of fermentation of chicha,a traditional beverage from Peru

Chicha is a drink prepared in several Andean countries from Inca's times by maize fermentation. Currently this fermentation is carried out in familiar artesanal “chicherías” that make one of the most known types of chicha, the “chicha de jora”. In this study we isolate and identify the yeasts mainly responsible of the fermentation process in this type of chicha in 10 traditional “chicherías” in Cusco region in Peru. We applied by first time MALDI-TOF MS analysis for the identification of yeast of non-clinic origin and the results showed that all of yeast strains isolated belong to the species Saccharomyces cerevisiae. These results agree with those obtained after the analysis of the D1/D2 and 5.8S-ITS regions. However the chicha strains have a phenotypic profile that differed in more than 40% as compared to that of current S. cerevisiae strains. To the best of our knowledge this is the first report concerning the yeasts involved in chicha fermentation.     

Mutations in Desmin's Carboxy-Terminal “Tail” Domain Severely Modify Filament and Network Mechanics

Inherited mutations in the gene coding for the intermediate filament protein desmin have been demonstrated to cause severe skeletal and cardiac myopathies. Unexpectedly, some of the mutated desmins, in particular those carrying single amino acid alterations in the non-α-helical carboxy-terminal domain (“tail”), have been demonstrated to form apparently normal filaments both in vitro and in transfected cells. Thus, it is not clear if filament properties are affected by these mutations at all. For this reason, we performed oscillatory shear experiments with six different desmin “tail” mutants in order to characterize the mesh size of filament networks and their strain stiffening properties. Moreover, we have carried out high-frequency oscillatory squeeze flow measurements to determine the bending stiffness of the respective filaments, characterized by the persistence length l_p. Interestingly, mesh size was not altered for the mutant filament networks, except for the mutant DesR454W, which apparently did not form proper filament networks. Also, the values for bending stiffness were in the same range for both the “tail” mutants (l_p = 1.0-2.0 μm) and the wild-type desmin (l_p = 1.1 ± 0.5 μm). However, most investigated desmin mutants exhibited a distinct reduction in strain stiffening compared to wild-type desmin and promoted nonaffine network deformation. Therefore, we conclude that the mutated amino acids affect intrafilamentous architecture and colloidal interactions along the filament in such a way that the response to applied strain is significantly altered.In order to explore the importance of the “tail” domain as such for filament network properties, we employed a “tail”-truncated desmin. Under standard conditions, it formed extended regular filaments, but failed to generate strain stiffening. Hence, these data strongly indicate that the “tail” domain is responsible for attractive filament-filament interactions. Moreover, these types of interactions may also be relevant to the network properties of the desmin cytoskeleton in patient muscle.     

Probing the metal ion selectivity in methionine aminopeptidase via changes in the luminescence properties of the enzyme bound europium ion

We report herein, for the first time, that Europium ion (Eu³⁺) binds to the “apo” form of Escherichia coli methionine aminopeptidase (EcMetAP), and such binding results in the activation of the enzyme as well as enhancement in the luminescence intensity of the metal ion. Due to competitive displacement of the enzyme-bound Eu³⁺ by different metal ions, we could determine the binding affinities of both “activating” and “non-activating” metal ions for the enzyme via fluorescence spectroscopy. The experimental data revealed that among all metal ions, Fe²⁺ exhibited the highest binding affinity for the enzyme, supporting the notion that it serves as the physiological metal ion for the enzyme. However, the enzyme-metal binding data did not adhere to the Irving-William series. On accounting for the binding affinity vis a vis the catalytic efficiency of the enzyme for different metal ions, it appears evident that that the “coordination states” and the relative softness” of metal ions are the major determinants in facilitating the EcMetAP catalyzed reaction.     

SAM recognition and conformational switching mechanism in the Bacillus subtilis yitJ S box/SAM-I riboswitch

S-box (SAM-I) riboswitches are a widespread class of riboswitches involved in the regulation of sulfur metabolism in Gram-positive bacteria. We report here the 3.0-Å crystal structure of the aptamer domain of the Bacillus subtilis yitJ S-box (SAM-I) riboswitch bound to S-adenosyl-l-methionine (SAM). The RNA folds into two sets of helical stacks spatially arranged by tertiary interactions including a K-turn and a pseudoknot at a four-way junction. The tertiary structure is further stabilized by metal coordination, extensive ribose zipper interactions, and SAM-mediated tertiary interactions. Despite structural differences in the peripheral regions, the SAM-binding core of the B. subtilis yitJ riboswitch is virtually superimposable with the previously determined Thermoanaerobacter tengcongensis yitJ riboswitch structure, suggesting that a highly conserved ligand-recognition mechanism is utilized by all S-box riboswitches. SHAPE (selective 2′-hydroxyl acylation analyzed by primer extension) chemical probing analysis further revealed that the alternative base-pairing element in the expression platform controls the conformational switching process. In the absence of SAM, the apo yitJ aptamer domain folds predominantly into a pre-binding conformation that resembles, but is not identical with, the SAM-bound state. We propose that SAM enters the ligand-binding site through the “J1/2-J3/4” gate and “locks” down the SAM-bound conformation through an induced-fit mechanism. Temperature-dependent SHAPE revealed that the tertiary interaction-stabilized SAM-binding core is extremely stable, likely due to the cooperative RNA folding behavior. Mutational studies revealed that certain modifications in the SAM-binding region result in loss of SAM binding and constitutive termination, which suggests that these mutations lock the RNA into a form that resembles the SAM-bound form in the absence of SAM.     

Melanization in the genus Propithecus Malagasey Lemur

Among the Lemurs of Madagascar the genus Propithecus contributes two species, but these are differentiated into numerous forms (subspecies and intermediates between them) which are essentially characterized by pelage coloration. The existence of certain “melanizing” forms among these intermediates permits us to examine the problem of the evolution of this genus in Madagascar, invoking a hypothesis to explain these chromatic variations. Hershkovitz's theory of “Metachromism” could easily fit the case of Propithecus. It would permit derivation of the light-coloured forms of the north, west and the south from a primitive form close to the dark-coloured Propithecus diadema of the southeast.     

The crystal structure analysis of group B Streptococcus sortase C1: a model for the "lid" movement upon substrate binding

A unique feature of the class-C-type sortases, enzymes essential for Gram-positive pilus biogenesis, is the presence of a flexible “lid” anchored in the active site. However, the mechanistic details of the “lid” displacement, suggested to be a critical prelude for enzyme catalysis, are not yet known. This is partly due to the absence of enzyme-substrate and enzyme-inhibitor complex crystal structures. We have recently described the crystal structures of the Streptococcus agalactiae SAG2603 V/R sortase SrtC1 in two space groups (type II and type III) and that of its “lid” mutant and proposed a role of the “lid” as a protector of the active-site hydrophobic environment. Here, we report the crystal structures of SAG2603 V/R sortase C1 in a different space group (type I) and that of its complex with a small-molecule cysteine protease inhibitor. We observe that the catalytic Cys residue is covalently linked to the small-molecule inhibitor without lid displacement. However, the type I structure provides a view of the sortase SrtC1 lid displacement while having structural elements similar to a substrate sorting motif suitably positioned in the active site. We propose that these major conformational changes seen in the presence of a substrate mimic in the active site may represent universal features of class C sortase substrate recognition and enzyme activation.     

Two reaction pathways for transformation of high potential cytochrome b559 of PS II into the intermediate potential form

This study describes an analysis of different treatments that influence the relative content and the midpoint potential of HP Cyt b559 in PS II membrane fragments from higher plants. Two basically different types of irreversible modification effects are distinguished: the HP form of Cyt b559 is either predominantly affected when the heme group is oxidized (“O-type” effects) or when it is reduced (“R-type” effects). Transformation of HP Cyt b559 to lower potential redox forms (IP and LP forms) by the “O-type” mechanism is induced by high pH and detergent treatments. In this case the effects consist of a gradual decrease in the relative content of HP Cyt b559 while its midpoint potential remains unaffected. Transformation of HP Cyt b559 via an “R-type” mechanism is caused by a number of exogenous compounds denoted L: herbicides, ADRY reagents and tetraphenylboron. These compounds are postulated to bind to the PS II complex at a quinone binding site designated as Q_C which interacts with Cyt b559 and is clearly not the Q_B site. Binding of compounds L to the Q_C site when HP Cyt b559 is oxidized gives rise to a gradual decrease in the E_m of HP Cyt b559 with increasing concentration of L (up to 10 K_ox(L) values) while the relative content of HP Cyt b559 is unaffected. Higher concentrations of compounds L required for their binding to Q_C site when HP Cyt b559 is reduced (described by K_red(L)) induce a conversion of HP Cyt b559 to lower potential redox forms (“R-type” transformation). Two reaction pathways for transitions of Cyt b559 between the different protein conformations that are responsible for the HP and IP/LP redox forms are proposed and new insights into the functional regulation of Cyt b559 via the Q_C site are discussed.     

A Report of Two cases of Al-Awadi Raas-Rothschild Syndrome (AARRS) supporting that “Apparent” Phocomelia differentiates AARRS from Schinzel Phocomelia Syndrome (SPS)

Although there is a long list of syndromes with phocomelia, there are only two syndromes in which there is concurrent pelvic dysplasia and phocomelia: Al-Awadi–Raas-Rothschild syndrome (AARRS) and Schinzel phocomelia syndrome (SPS). Currently, there is a diagnostic confusion between the two syndromes and both have the same MIM entry (MIM 276820). We believe that the two syndromes are different entities and we also believe that the limb defect in SPS is a “true” phocomelia while the limb defect in AARRS is an “apparent” phocomelia. “Apparent” phocomelia describes the most severe form of ulnar ray deficiency in which there is absent ulna with radio-humeral synostosis. “Apparent” phocomelia is diagnosed radiologically by three radiological features: the apparently single bone occupying the arm/forearm appears relatively long, the area of radio-humeral synostosis will have thicker cortex with or without slight angulation, and the lower end of the bone resembles the lower end of a radius and not a humerus. In this paper, we present two new cases of AARRS from two different Saudi Arabian tribes: one case with R292C mutation of WNT7A with bilateral “apparent” phocomelia and a second case with a novel c.814G>T mutation of the WNT7A gene (resulting in wnt7a protein truncation at position 272) with unilateral “apparent” phocomelia. We reviewed previously reported cases of AARRS and SPS to further delineate the differences between these two syndromes. We make the argument that these two syndromes are two different entities and hence require two different MIM entries.     

Antiparallel Conformation of Knob and Hole Aglycosylated Half-Antibody Homodimers Is Mediated by a CH2–CH3 Hydrophobic Interaction

Bispecific antibody and antibody-like molecules are of wide interest as potential therapeutics that can recognize two distinct targets. Among the variety of ways such molecules have been engineered is by creating “knob” and “hole” heterodimerization sites in the CH3 domains of two antibody heavy chains. The molecules produced in this manner maintain their biological activities while differing very little from the native human IgG sequence. To better understand the knob-into-hole interface, the molecular mechanism of heterodimerization, and to engineer Fc domains that could improve the assembly and purity of heterodimeric reaction products, we sought crystal structures of aglycosylated heterodimeric and homodimeric “knob” and “hole” Fc fragments derived from bacterial expression. The structure of the knob-into-hole Fc was determined at 2.64 Å. Except for the sites of mutation, the structure is very similar to that of the native human IgG1 Fc, consistent with a heterodimer interaction kinetic K_D of < 1 nM. Homodimers of the “knob” and “hole” mutants were also obtained, and their X-ray structures were determined at resolutions 2.5 Å and 2.1 Å, respectively. Both kinds of homodimers adopt a head-to-tail quaternary structure and thus do not contain direct knob/knob or hole/hole CH3 interactions. The head-to-tail arrangement was disfavored by adding site-directed mutations at F241 and F243 in the CH2 domains, leading to increases in both rate and efficiency of bispecific (heterodimer) assembly.