Harnessing and engineering amide bond forming ligases for the synthesis of amides

The amide functional group is ubiquitous in nature and one of the most important motifs in pharmaceuticals, agrochemicals, and other valuable products. While coupling amides and carboxylic acids is a trivial synthetic transformation, it often requires protective group manipulation, along with stoichiometric quantities of expensive and deleterious coupling reagents. Nature has evolved a range of enzymes to construct amide bonds, the vast majority of which utilize adenosine triphosphate to activate the carboxylic acid substrate for amine coupling. Despite the fact that these enzymes operate under mild conditions, as well as possessing chemoselectivity and regioselectivity that obviates the need for protecting groups, their synthetic potential has been largely unexplored. In this review, we discuss recent research into the discovery, characterization, and development of amide bond forming enzymes, with an emphasis on stand-alone ligase enzymes that can generate amides directly from simple carboxylic acid and amine substrates.     

Oreopithecus bambolii: an unlikely case of hominid-like grip capability in a Miocene ape

Oreopithecus bambolii, an ape from the late Miocene of Italy, is said to possess a hand capable of a precision grip like that of humans. Relative hand length, proportions of the thumb, and morphological features of the thumb and wrist were adduced to support the idea that Oreopithecus had a hand that closely matched the pattern in Australopithecus. A reappraisal of earlier arguments and comparisons of Oreopithecus with humans, apes, and Old World monkeys, reveals that Oreopithecus had an essentially ape-like hand that emphasized ape-like power grasping over human-like precision grasping.     

Changes in the spectral power and coherence of the EEG alpha rhythm in humans performing grasp efforts by the right and left arm

We compared changes in the EEG indices in healthy dextral volunteers performing static force grasps by the arm. Three test modes were used: (i) performance of two successive grasps by the dominant (right) arm (test A), (ii) performance of two successive grasps by the subdominant (left) arm (test B), and (iii) performance of the grasps first by the right arm and then by the left arm (test C). Fourteen, six, and nine persons took part in tests A–C, respectively. In the course of grasps performed by the right and left arms, bilateral increases in synchronization within the alpha 1 and alpha 2 ranges were frequently observed in occipital regions in both the first and repeated grasps (P < 0.05). Consecutive grasps by the right arm were accompanied by clear desynchronization in a few anterior and central leads. Alpha 2 desynchronization was observed in both realizations of the left-arm grasps (test B) performed by some subjects, but intragroup modifications were not significant in this case. The coherence coefficients of the alpha 2 rhythm in most cases increased for symmetric leads from the right and left hemispheres in the course of grasps by both the right and left hands. The effect of intensification of interhemisphere links was manifested in the anterior and central cortical regions; this fact showed that interhemisphere interaction increases in the course of the static effort. Changes in the coherence coefficients for the alpha 2 range in the performance of the grasp efforts by the right arm and the left arm were most clear in the posterotemporal (P = 0.02), parietal (P = 0.05), and anterofrontal (P = 0.06) lead pairs. Thus, we demonstrated the dependence between the side of performance of the muscle effort in the mode close to isometric and lateralization of the EEG modifications. Neirofiziologiya/Neurophysiology, Vol. 38, No. 3, pp. 235–238, May–June, 2006.     

The evolution of host‐specific variation in cuckoo eggshell strength

Cuckoo eggs are renowned for their mimicry of different host species, leading to the evolution of host‐specific races (or ‘gentes’) defined by egg colour and pattern. This study aims to test the prediction that another property of parasitic eggs, namely shell strength, might also have experienced divergent selection within cuckoo species. Host races of the common cuckoo Cuculus canorus encountering stronger host rejection have thicker‐shelled eggs than those parasitising less discriminating species, as expected if egg strengthening discourages host rejection. Moreover, in the diederik cuckoo Chrysococcyx caprius, eggshell thickness was correlated across cuckoo gentes and host species, as expected if eggshell strength has been involved in coevolutionary interactions. This is the first report of host‐specific differences in cuckoo egg properties other than colour and pattern and lends correlational support to the hypothesis that the strong eggshells of brood parasites are an adaptation to reduce host rejection.     

E2-binding surface on Uba3 β-grasp domain undergoes a conformational transition

The covalent attachment of ubiquitin (Ub) and ubiquitin‐like (Ubl) proteins to various eukaryotic targets plays critical roles in regulating numerous cellular processes. E1‐activating enzymes are critical, because they catalyze activation of their cognate Ub/Ubl protein and are responsible for its transfer to the correct E2‐conjugating enzyme(s). The activating enzyme for neural‐precursor‐cell‐expressed developmentally downregulated 8 (NEDD8) is a heterodimer composed of APPBP1 and Uba3 subunits. The carboxyl terminal ubiquitin‐like β‐grasp domain of human Uba3 (Uba3‐βGD) has been suggested as a key E2‐binding site defining E2 specificity. In crystal structures of free E1 and the NEDD8‐E1 complex, the E2‐binding surface on the domain was missing from the electron density. However, when complexed with various E2s, this missing segment adopts a kinked α‐helix. Here, we demonstrate that Uba3‐βGD is an independently folded domain in solution and that residues involved in E2 binding are absent from the NMR spectrum, indicating that the E2‐binding surface on Uba3‐βGD interconverts between multiple conformations, analogous to a similar conformational transition observed in the E2‐binding surface of SUMO E1. These results suggest that access to multiple conformational substates is an important feature of the E1–E2 interaction. Proteins 2012;. © 2012 Wiley Periodicals, Inc.     

Corticospinal mirror neurons

Here, we report the properties of neurons with mirror-like characteristics that were identified as pyramidal tract neurons (PTNs) and recorded in the ventral premotor cortex (area F5) and primary motor cortex (M1) of three macaque monkeys. We analysed the neurons’ discharge while the monkeys performed active grasp of either food or an object, and also while they observed an experimenter carrying out a similar range of grasps. A considerable proportion of tested PTNs showed clear mirror-like properties (52% F5 and 58% M1). Some PTNs exhibited ‘classical’ mirror neuron properties, increasing activity for both execution and observation, while others decreased their discharge during observation (‘suppression mirror-neurons’). These experiments not only demonstrate the existence of PTNs as mirror neurons in M1, but also reveal some interesting differences between M1 and F5 mirror PTNs. Although observation-related changes in the discharge of PTNs must reach the spinal cord and will include some direct projections to motoneurons supplying grasping muscles, there was no EMG activity in these muscles during action observation. We suggest that the mirror neuron system is involved in the withholding of unwanted movement during action observation. Mirror neurons are differentially recruited in the behaviour that switches rapidly between making your own movements and observing those of others.     

Grasp modelling with a biomechanical model of the hand

The use of a biomechanical model for human grasp modelling is presented. A previously validated biomechanical model of the hand has been used. The equilibrium of the grasped object was added to the model through the consideration of a soft contact model. A grasping posture generation algorithm was also incorporated into the model. All the geometry was represented using a spherical extension of polytopes (s-topes) for efficient collision detection. The model was used to simulate an experiment in which a subject was asked to grasp two cylinders of different diameters and weights. Different objective functions were checked to solve the indeterminate problem. The normal finger forces estimated by the model were compared to those experimentally measured. The popular objective function sum of the squared muscle stresses was shown not suitable for the grasping simulation, requiring at least being complemented by task-dependent grasp quality measures.     

Structure and function of Escherichia coli RimK,an ATP‐grasp fold,l‐glutamyl ligase enzyme

We report herein the crystal structure of Escherichia coli RimK at a resolution of 2.85 Å, an enzyme that catalyzes the post‐translational addition of up to 15 C‐terminal glutamate residues to ribosomal protein S6. The structure belongs to the ATP‐grasp superfamily and is organized as a tetramer, consistent with gel filtration analysis. Each subunit consists of three distinct structural domains and the active site is located in the cleft between these domains. The catalytic reaction appears to occur at the junction between the three domains as ATP binds between the B and C domains, and other substrates bind nearby.Proteins 2013; 81:1847–1854. © 2013 Wiley Periodicals, Inc.