NlpC/P60 domain-containing proteins of Mycobacterium avium subspecies paratuberculosis that differentially bind and hydrolyze peptidoglycan

A subset of proteins containing NlpC/P60 domains are bacterial peptidoglycan hydrolases that cleave noncanonical peptide linkages and contribute to cell wall remodeling as well as cell separation during late stages of division. Some of these proteins have been shown to cleave peptidoglycan in Mycobacterium tuberculosis and play a role in Mycobacterium marinum virulence of zebra fish; however, there are still significant knowledge gaps concerning the molecular function of these proteins in Mycobacterium avium subspecies paratuberculosis (MAP). The MAP genome sequence encodes five NlpC/P60 domain-containing proteins. We describe atomic resolution crystal structures of two such MAP proteins, MAP_1272c and MAP_1204. These crystal structures, combined with functional assays to measure peptidoglycan cleavage activity, led to the observation that MAP_1272c does not have a functional catalytic core for peptidoglycan hydrolysis. Furthermore, the structure and sequence of MAP_1272c demonstrate that the catalytic residues normally required for hydrolysis are absent, and the protein does not bind peptidoglycan as efficiently as MAP_1204. While the NlpC/P60 catalytic triad is present in MAP_1204, changing the catalytic cysteine-155 residue to a serine significantly diminished catalytic activity, but did not affect binding to peptidoglycan. Collectively, these findings suggest a broader functional repertoire for NlpC/P60 domain-containing proteins than simply hydrolases

Avian thermoregulation in the heat : evaporative cooling capacity of arid-zone Caprimulgiformes from two continents

Birds in the order Caprimulgiformes (nightjars and allies) have a remarkable capacity for thermoregulation over a wide range of environmental temperatures, exhibiting pronounced heterothermy in cool conditions and extreme heat tolerance at high environmental temperatures. We measured thermoregulatory responses to acute heat stress in three species of Caprimulgiformes that nest in areas of extreme heat and aridity, the common poorwill (Phalaenoptilus nuttallii: Caprimulgidae) and lesser nighthawk (Chordeiles acutipennis: Caprimulgidae) in the Sonoran Desert of Arizona, and the Australian owlet-nightjar (Aegotheles cristatus: Aegothelidae) in the mallee woodlands of South Australia. We exposed wild-caught birds to progressively increasing air temperatures (Ta) and measured resting metabolic rate (RMR), evaporative water loss (EWL), body temperature (Tb) and heat tolerance limit (HTL; the maximum Ta reached). Comparatively low RMR values were observed in all species (0.35, 0.36 and 0.40 Wfor the poorwill, nighthawk and owletnightjar, respectively), with Tb approximating Ta at 40°C and mild hyperthermia occurring as Ta reached the HTL. Nighthawks and poorwills reached HTLs of 60 and 62°C, respectively, whereas the owlet-nightjar had a HTL of 52°C. RMR increased gradually above minima at Ta of 42, 42 and 35°C, and reached 1.7, 1.9 and 2.0 times minimum resting values at HTLs in the poorwill, nighthawk and owletnightjar, respectively. EWL increased rapidly and linearly as Ta exceeded Tb and resulted in maximum rates of evaporative heat dissipation equivalent to 237–424% of metabolic heat production. Bouts of gular flutter resulted in large transient increases in evaporative heat loss (50–123%) accompanied by only small increments in RMR (<5%). The cavity-nesting/roosting owletnightjar had a lower HTL and less efficient evaporative cooling compared with the species that nest and/or roost on open desert surfaces. The high efficiency of gular flutter for evaporative cooling, combined with mild hyperthermia, provides the physiological basis for defending Tb well below Ta in extreme heat and is comparable to the efficient cooling observed in arid-zone columbids in which cutaneous EWL is the predominant cooling pathway.The National Science Foundation under IOS-1122228 to B.O.W.http://jeb.biologists.org2018-10-01am2018Zoology and Entomolog

Avian thermoregulation in the heat : evaporative cooling in five Australian passerines reveals within-order biogeographic variation in heat tolerance

Evaporative heat loss pathways vary among avian orders, but the extent to which evaporative cooling capacity and heat tolerance vary within orders remains unclear. We quantified the upper limits to thermoregulation under extremely hot conditions in five Australian passerines: yellow-plumed honeyeater (Lichenostomus ornatus; ∼17 g), spiny-cheeked honeyeater (Acanthagenys rufogularis; ∼42 g), chestnut-crowned babbler (Pomatostomus ruficeps; ∼52 g), grey butcherbird (Cracticus torquatus; ∼86 g) and apostlebird (Struthidea cinerea; ∼118 g). At air temperatures (Ta) exceeding body temperature (Tb), all five species showed increases in Tb to maximum values around 44–45°C, accompanied by rapid increases in resting metabolic rate above clearly defined upper critical limits of thermoneutrality and increases in evaporative water loss (EWL) to levels equivalent to 670–860% of baseline rates at thermoneutral Ta. Maximum cooling capacity, quantified as the fraction of metabolic heat production dissipated evaporatively, ranged from 1.20 to 2.17, consistent with the known range for passerines, and well below the corresponding ranges for columbids and caprimulgids. Heat tolerance limit (HTL, the maximum Ta tolerated) scaled positively with body mass, varying from 46°C in yellow-plumed honeyeaters to 52°C in a single apostlebird, but was lower than that of three southern African ploceid passerines investigated previously. We argue this difference is functionally linked to a smaller scope for increases in EWL above baseline levels. Our data reiterate the reliance of passerines in general on respiratory evaporative heat loss via panting, but also reveal substantial within-order variation in heat tolerance and evaporative cooling capacity.The National Science Foundation [IOS- 1122228 to B.O.W.].http://jeb.biologists.org2018-07-30am2017Zoology and Entomolog

A Molecular Insight into Complement Evasion by the Staphylococcal Complement Inhibitor Protein Family

Staphylococcus aureus possesses an impressive arsenal of complement evasion proteins that help the bacterium escape attack of the immune system. The staphylococcal complement inhibitor (SCIN) protein exhibits a particularly high potency and was previously shown to block complement by acting at the level of the C3 convertases. However, many details about the exact binding and inhibitory mechanism remained unclear. In this study, we demonstrate that SCIN directly binds with nanomolar affinity to a functionally important area of C3b that lies near the C terminus of its -chain. Direct competition of SCIN with factor B for C3b slightly decreased the formation of surface-bound convertase. However, the main inhibitory effect can be attributed to an entrapment of the assembled convertase in an inactive state. Whereas native C3 is still able to bind to the blocked convertase, no generation and deposition of C3b could be detected in the presence of SCIN. Furthermore, SCIN strongly competes with the binding of factor H to C3b and influences its regulatory activities: the SCIN-stabilized convertase was essentially insensitive to decay acceleration by factor H and the factor I- and H-mediated conversion of surface-bound C3b to iC3b was significantly reduced. By targeting a key area on C3b, SCIN is able to block several essential functions within the alternative pathway, which explains the high potency of the inhibitor. Our findings provide an important insight into complement evasion strategies by S. aureus and may act as a base for further functional studies. The Journal of Immunology, 2009, 183: 2565–2574. T he bacterium Staphylococcus aureus is a widely dissem-inated and persistent human pathogen that has a long-standing and increasingly negative impact on human health. It is a primary etiologic agent of numerous disorders tha

Introductory programming: a systematic literature review

As computing becomes a mainstream discipline embedded in the school curriculum and acts as an enabler for an increasing range of academic disciplines in higher education, the literature on introductory programming is growing. Although there have been several reviews that focus on specific aspects of introductory programming, there has been no broad overview of the literature exploring recent trends across the breadth of introductory programming. This paper is the report of an ITiCSE working group that conducted a systematic review in order to gain an overview of the introductory programming literature. Partitioning the literature into papers addressing the student, teaching, the curriculum, and assessment, we explore trends, highlight advances in knowledge over the past 15 years, and indicate possible directions for future research

Paracellular absorption is relatively low in the herbivorous Egyptian spiny-tailed lizard, Uromastyx aegyptia

Extent: 9 p.Absorption of small water-soluble nutrients in vertebrate intestines occurs both by specific, mediated transport and by nonspecific, passive, paracellular transport. Although it is apparent that paracellular absorption represents a significant route for nutrient absorption in many birds and mammals, especially small, flying species, its importance in ectothermic vertebrates has not previously been explored. Therefore, we measured fractional absorption (e) and absorption rate of three paracellular probes (arabinose, L-rhamnose, cellobiose) and of 3-O-methyl D-glucose (absorbed by both mediated and paracellular pathways) by the large herbivorous lizard, Uromastyx aegyptia, to explore the relative importance of paracellular and mediated transport in an ectothermic, terrestrial vertebrate. Fractional absorption of 3-O-methyl D-glucose was high (e = 0.7360.04) and similar to other vertebrates; e of the paracellular probes was relatively low (arabinose e = 0.3160.03, Lrhamnose e = 0.1960.02, and cellobiose e = 0.1460.02), and decreased with molecular mass, a pattern consistent with other vertebrates. Paracellular absorption accounted for approximately 24% of total 3-O-methyl D-glucose uptake, indicating low reliance on this pathway for these herbivorous lizards, a pattern similar to that found in other terrestrial vertebrates, and different from small flying endotherms (both birds and bats).Todd J. McWhorter, Berry Pinshow, William H. Karasov and Christopher R. Trac

NlpC/P60 domain-containing proteins of Mycobacterium avium subspecies paratuberculosis that differentially bind and hydrolyze peptidoglycan

A subset of proteins containing NlpC/P60 domains are bacterial peptidoglycan hydrolases that cleave noncanonical peptide linkages and contribute to cell wall remodeling as well as cell separation during late stages of division. Some of these proteins have been shown to cleave peptidoglycan in Mycobacterium tuberculosis and play a role in Mycobacterium marinum virulence of zebra fish; however, there are still significant knowledge gaps concerning the molecular function of these proteins in Mycobacterium avium subspecies paratuberculosis (MAP). The MAP genome sequence encodes five NlpC/P60 domain-containing proteins. We describe atomic resolution crystal structures of two such MAP proteins, MAP_1272c and MAP_1204. These crystal structures, combined with functional assays to measure peptidoglycan cleavage activity, led to the observation that MAP_1272c does not have a functional catalytic core for peptidoglycan hydrolysis. Furthermore, the structure and sequence of MAP_1272c demonstrate that the catalytic residues normally required for hydrolysis are absent, and the protein does not bind peptidoglycan as efficiently as MAP_1204. While the NlpC/P60 catalytic triad is present in MAP_1204, changing the catalytic cysteine-155 residue to a serine significantly diminished catalytic activity, but did not affect binding to peptidoglycan. Collectively, these findings suggest a broader functional repertoire for NlpC/P60 domain-containing proteins than simply hydrolases

Hummingbirds rely on both paracellular and carrier-mediated intestinal glucose absorption to fuel high metabolism

Twenty years ago, the highest active glucose transport rate and lowest passive glucose permeability in vertebrates were reported in Rufous and Anna's hummingbirds (Selasphorus rufus, Calypte anna). These first measurements of intestinal nutrient absorption in nectarivores provided an unprecedented physiological foundation for understanding their foraging ecology. They showed that physiological processes are determinants of feeding behaviour. The conclusion that active, mediated transport accounts for essentially all glucose absorption in hummingbirds influenced two decades of subsequent research on the digestive physiology and nutritional ecology of nectarivores. Here, we report new findings demonstrating that the passive permeability of hummingbird intestines to glucose is much higher than previously reported, suggesting that not all sugar uptake is mediated. Even while possessing the highest active glucose transport rates measured in vertebrates, hummingbirds must rely partially on passive non-mediated intestinal nutrient absorption to meet their high mass-specific metabolic demands