Impacts of land-use management on ecosystem services and biodiversity: an agent-based modelling approach.

The science of ecosystem service (ES) mapping has become increasingly sophisticated over the past 20 years, and examples of successfully integrating ES into management decisions at national and sub-national scales have begun to emerge. However, increasing model sophistication and accuracy-and therefore complexity-may trade-off with ease of use and applicability to real-world decision-making contexts, so it is vital to incorporate the lessons learned from implementation efforts into new model development. Using successful implementation efforts for guidance, we developed an integrated ES modelling system to quantify several ecosystem services: forest timber production and carbon storage, water purification, pollination, and biodiversity. The system is designed to facilitate uptake of ES information into land-use decisions through three principal considerations: (1) using relatively straightforward models that can be readily deployed and interpreted without specialized expertise; (2) using an agent-based modelling framework to enable the incorporation of human decision-making directly within the model; and (3) integration among all ES models to simultaneously demonstrate the effects of a single land-use decision on multiple ES. We present an implementation of the model for a major watershed in Alberta, Canada, and highlight the system's capabilities to assess a suite of ES under future management decisions, including forestry activities under two alternative timber harvest strategies, and through a scenario modelling analysis exploring different intensities of hypothetical agricultural expansion. By using a modular approach, the modelling system can be readily expanded to evaluate additional ecosystem services or management questions of interest in order to guide land-use decisions to achieve socioeconomic and environmental objectives.

Catalase (KatA) plays a role in protection against anaerobic nitric oxide in Pseudomonas aeruginosa.

Pseudomonas aeruginosa (PA) is a common bacterial pathogen, responsible for a high incidence of nosocomial and respiratory infections. KatA is the major catalase of PA that detoxifies hydrogen peroxide (H2O2), a reactive oxygen intermediate generated during aerobic respiration. Paradoxically, PA displays elevated KatA activity under anaerobic growth conditions where the substrate of KatA, H2O2, is not produced. The aim of the present study is to elucidate the mechanism underlying this phenomenon and define the role of KatA in PA during anaerobiosis using genetic, biochemical and biophysical approaches. We demonstrated that anaerobic wild-type PAO1 cells yielded higher levels of katA transcription and expression than aerobic cells, whereas a nitrite reductase mutant ΔnirS produced ∼50% the KatA activity of PAO1, suggesting that a basal NO level was required for the increased KatA activity. We also found that transcription of the katA gene was controlled, in part, by the master anaerobic regulator, ANR. A ΔkatA mutant and a mucoid mucA22 ΔkatA bacteria demonstrated increased sensitivity to acidified nitrite (an NO generator) in anaerobic planktonic and biofilm cultures. EPR spectra of anaerobic bacteria showed that levels of dinitrosyl iron complexes (DNIC), indicators of NO stress, were increased significantly in the ΔkatA mutant, and dramatically in a ΔnorCB mutant compared to basal levels of DNIC in PAO1 and ΔnirS mutant. Expression of KatA dramatically reduced the DNIC levels in ΔnorCB mutant. We further revealed direct NO-KatA interactions in vitro using EPR, optical spectroscopy and X-ray crystallography. KatA has a 5-coordinate high spin ferric heme that binds NO without prior reduction of the heme iron (Kd ∼6 µM). Collectively, we conclude that KatA is expressed to protect PA against NO generated during anaerobic respiration. We proposed that such protective effects of KatA may involve buffering of free NO when potentially toxic concentrations of NO are approached.

Structural basis for Zn2+-dependent intercellular adhesion in staphylococcal biofilms.

Staphylococcal bacteria, including Staphylococcus epidermidis and Staphylococcus aureus, cause chronic biofilm-related infections. The homologous proteins Aap and SasG mediate biofilm formation in S. epidermidis and S. aureus, respectively. The self-association of these proteins in the presence of Zn(2+) leads to the formation of extensive adhesive contacts between cells. This study reports the crystal structure of a Zn(2+) -bound construct from the self-associating region of Aap. Several unusual structural features include elongated ß-sheets that are solvent-exposed on both faces and the lack of a canonical hydrophobic core. Zn(2+)-dependent dimers are observed in three distinct crystal forms, formed via pleomorphic coordination of Zn(2+) in trans across the dimer interface. These structures illustrate how a long, flexible surface protein is able to form tight intercellular adhesion sites under adverse environmental conditions.

Structural comparison of ColH and ColG collagen-binding domains from Clostridium histolyticum.

Clostridium histolyticum secretes collagenases, ColG and ColH, that cause extensive tissue destruction in myonecrosis. The C-terminal collagen-binding domain (CBD) of collagenase is required for insoluble collagen fibril binding and subsequent collagenolysis. The high-resolution crystal structures of ColG-CBD (s3b) and ColH-CBD (s3) are reported in this paper. The new X-ray structure of s3 was solved at 2.0-Å resolution (R = 17.4%; R(free) = 23.3%), while the resolution of the previously determined s3b was extended to 1.4 Å (R = 17.9%; R(free) = 21.0%). Despite sharing only 30% sequence identity, the molecules resemble one another closely (root mean square deviation [RMSD] C(α) = 1.5 Å). All but one residue, whose side chain chelates with Ca(2+), are conserved. The dual Ca(2+) binding site in s3 is completed by an unconserved aspartate. Differential scanning calorimetric measurements showed that s3 gains thermal stability, comparable to s3b, by binding to Ca(2+) (holo T(m) = 94.1°C; apo T(m) = 70.2°C). holo s3 is also stabilized against chemical denaturants urea and guanidine HCl. The three most critical residues for collagen interaction in s3b are conserved in s3. The general shape of the binding pocket is retained by altered loop structures and side chain positions. Small-angle X-ray scattering data revealed that s3 also binds asymmetrically to minicollagen. Besides the calcium-binding sites and the collagen-binding pocket, architecturally important hydrophobic residues and the hydrogen-bonding network around the cis-peptide bond are well conserved within the metallopeptidase subfamily M9B. CBDs were previously shown to bind to the extracellular matrix of various tissues. Compactness and extreme stability in physiological Ca(2+) concentration possibly make both CBDs suitable for targeted growth factor delivery.

Infrared multiphoton dissociation in quadrupole ion traps.

The development of new ion activation techniques continues to be a dynamic area of scientific discovery, in part to complement the tremendous innovations in ionization methods that have allowed the mass spectrometric analysis of an enormous array of molecules. Ion activation/dissociation provides key information about ion structures, binding energies, and differentiation of isomers, as well as affording a primary means of identifying compounds in mixtures. Numerous new activation methods have emerged over the past two decades in an effort to develop alternatives to collisional activated dissociation, the gold standard for providing structurally diagnostic fragmentation patterns. Collisional activated dissociation does not always offer sufficiently high or controllable energy deposition, thus rendering it less useful for certain classes of molecules, such as large proteins or macromolecular complexes. Photodissociation is one of the most promising alternatives and is readily implemented in ion trapping and time-of-flight mass spectrometers. Photodissociation generally entails using a laser to irradiate ions with UV, visible, or IR photons, thus resulting in internal energy deposition based on the number and wavelengths of the photons. The activation process can be extremely rapid and efficient, as well as having the potential for high total energy deposition. This review describes infrared multiphoton dissociation in quadrupole ion trap mass spectrometry. A comparison of photodissociation and collisional activated dissociation is covered, in addition to some of the methods to increase photodissociation efficiency. Numerous applications of IRMPD are discussed as well, including ones related to the analysis of drugs, peptides, nucleic acids, and oligosaccharides.

Improved infrared multiphoton dissociation of peptides through N-terminal phosphonite derivatization.

A strategy for improving the sequencing of peptides by infrared multiphoton dissociation (IRMPD) in a linear ion trap mass spectrometer is described. We have developed an N-terminal derivatization reagent, 4-methylphosphonophenylisothiocyanate (PPITC), which allows the attachment of an IR-chromogenic phosphonite group to the N-terminus of peptides, thus enhancing their IRMPD efficiencies. After the facile derivatization process, the PPITC-modified peptides require shorter irradiation times for efficient IRMPD and yield extensive series of y ions, including those of low m/z that are not detected upon traditional CID. The resulting IRMPD mass spectra afford more complete sequence coverage for both model peptides and tryptic peptides from cytochrome c. We compare the effectiveness of this derivatization/IRMPD approach to that of a common N-terminal sulfonation reaction that utilizes 4-sulfophenylisothiocyanate (SPITC) in conjunction with CID and IRMPD.

Ultraviolet photodissociation at 355 nm of fluorescently labeled oligosaccharides.

Ultraviolet photodissociation (UVPD) produces complementary fragmentation to collision-induced dissociation (CID) when implemented for activation of fluorescently labeled oligosaccharide and glycan ions. Reductive amination of oligosaccharides with fluorophore reagents results in efficient photon absorption at 355 nm, producing fragment ions from the nonreducing end that do not contain the appended fluorophore. In contrast to the fragment ions observed upon UVPD (A- and C-type ions), CID produces mainly reducing end fragments retaining the fluorophore (Y-type ions). UVPD affords better isomeric differentiation of both the lacto-N-fucopentaoses series and the lacto-N-difucohexaoses series, but in general, the combination of UVPD and CID offers the most diagnostic elucidation of complex branched oligosaccharides. Four fluorophores yielded similar MS/MS results; however, 6-aminoquinoline (6-AQ), 2-amino-9(10 H)-acridone (AMAC) and 7-aminomethylcoumarin (AMC) afforded more efficient photon absorption and subsequent dissociation than 2-aminobenzamide (2-AB). UVPD also was useful for characterization of glycans released from ribonuclease B and derivatized with 6-AQ. Lastly, electron photodetachment dissociation of oligosaccharides derivatized with 7-amino-1,3-naphthalenedisulfonic acid (AGA) yielded unique cross-ring cleavages similar to those obtained by electron detachment dissociation.

RAM-induced allostery facilitates assembly of a notch pathway active transcription complex.

The Notch pathway is a conserved cell-to-cell signaling mechanism, in which extracellular signals are transduced into transcriptional outputs through the nuclear effector CSL. CSL is converted from a repressor to an activator through the formation of the CSL-NotchIC-Mastermind ternary complex. The RAM (RBP-J associated molecule) domain of NotchIC avidly interacts with CSL; however, its role in assembly of the CSL-NotchIC-Mastermind ternary complex is not understood. Here we provide a comprehensive thermodynamic, structural, and biochemical analysis of the RAM-CSL interaction for components from both mouse and worm. Our binding data show that RAM and CSL form a high affinity complex in the presence or absence of DNA. Our structural studies reveal a striking distal conformational change in CSL upon RAM binding, which creates a docking site for Mastermind to bind to the complex. Finally, we show that the addition of a RAM peptide in trans facilitates formation of the CSL-NotchIC-Mastermind ternary complex in vitro.

Photodissociation of non-covalent peptide-crown ether complexes.

Highly chromogenic 18-crown-6-dipyrrolylquinoxaline coordinates primary amines of peptides, forming non-covalent complexes that can be transferred to the gas-phase by electrospray ionization. The appended chromogenic crown ether facilitates efficient energy transfer to the peptide upon ultraviolet irradiation in the gas phase, resulting in diagnostic peptide fragmentation. Collisional-activated dissociation and infrared multiphoton dissociation of these non-covalent complexes result only in their disassembly with the charge retained on either the peptide or crown ether, yielding no sequence ions. Upon UV photon absorption the intermolecular energy transfer is facilitated by the fast activation timescale of ultraviolet photodissociation (<10 ns) and by the collectively strong hydrogen bonding between the crown ether and peptide, thus allowing effective transfer of energy to the peptide moiety before disruption of the intermolecular hydrogen bonds.

MS/MS simplification by 355 nm ultraviolet photodissociation of chromophore-derivatized peptides in a quadrupole ion trap.

Ultraviolet photodissociation (UVPD) of chromophore-modified peptides enhances the capabilities for de novo sequencing in a quadrupole ion trap mass spectrometer. Attachment of UV chromophores allows efficient photoactivation of not only the precursor ions but also any fragments that retain the chromophore functionality. For doubly protonated peptides, UVPD leads to a vast reduction in MS/MS complexity. The array of b and y ions typically seen upon collisionally activated dissociation is reduced to a single series of either y or b ions by UVPD depending on the location of the chromophore (i.e., N- or C-terminus). The sulfonation reagent Alexa Fluor 350 (AF350) provided the best overall results for the singly and doubly charged peptides by UVPD. The nonsulfonated analogue of AF350, 7-amino-4-methylcoumarin-3-acetic acid, also led to simplified spectra for doubly charged, but not singly charged, peptides by UVPD. Dinitrophenyl-peptides also yielded simplified spectra by UVPD albeit with a small amount of internal fragments accompanying the series of diagnostic y ions. The success of this MS/MS simplification process stems from extensive secondary fragmentation of any chromophore-containing fragments upon exposure to subsequent laser pulses. Energy-variable UVPD reveals that the abundances of non-chromophore-containing y fragment ions increase linearly with laser pulse energy, suggesting secondary dissociation of these species is insignificant. The abundances of chromophore-containing a/b fragment ions follow a quadratic trend due to the extensive secondary fragmentation at higher laser energies or multiple pulses.

ADHD and substance use disorders: developmental aspects and the impact of stimulant treatment.

Adolescents and adults with substance use disorders often demonstrate symptoms of inattention, impulsivity, and hyperactivity. These core symptoms of ADHD may contribute to the development of substance use disorders by promoting antisocial behavior and substance use; conversely, substance use itself can adversely affect these symptoms. Common deficits in self-regulatory processes could underlie the developmental progression of these disorders, deficits further worsened by ongoing substance use. Some investigators have questioned whether stimulant treatment itself could promote substance abuse, while others have argued that such treatment reduces substance abuse. With an increased awareness of the phenomenon of adult ADHD and its relevance to substance-abusing persons, there is an increased awareness of the potential benefit of ADHD treatment on substance abuse treatment outcome. Consideration of an individual's developmental relationship between attention deficit/hyperactivity symptoms and substance use can inform treatment planning among patients seeking substance abuse treatment.

Infrared multiphoton dissociation of duplex DNA/drug complexes in a quadrupole ion trap.

Noncovalent duplex DNA/drug complexes formed between one of three 14-base pair non-self-complementary duplexes with variable GC content and one of eight different DNA-interactive drugs are characterized by infrared multiphoton dissociation (IRMPD), and the resulting spectra are compared to conventional collisionally activated dissociation (CAD) mass spectra in a quadrupole ion trap mass spectrometer. IRMPD yielded comparable information to previously reported CAD results in which strand separation pathways dominate for complexes containing the more AT-rich sequences and/or minor groove binding drugs, whereas drug ejection pathways are prominent for complexes containing intercalating drugs and/or duplexes with higher GC base content. The large photoabsorptive cross section of the phosphate backbone at 10.6 mum promotes highly efficient dissociation within short irradiation times (<2 ms at 50 W) or using lower laser powers and longer irradiation times (<15 W at 15 ms), activation times on par with or shorter than standard CAD experiments. This large photoabsorptivity leads to a controllable ion activation method which can be used to produce qualitatively similar spectra to CAD while minimizing uninformative base loss dissociation pathways or instead be tuned to yield a high degree of secondary fragmentation. Additionally, the low-mass cutoff associated with conventional CAD plays no role in IRMPD, resulting in richer MS/MS information in the low m/z region. IRMPD is also used for multiadduct dissociation in order to increase MS/MS sensitivity, and a two-stage IRMPD/IRMPD method is demonstrated as a means to give specific DNA sequence information that would be useful when screening drug binding by mixtures of duplexes.

Amplification of infrared multiphoton dissociation efficiency in a quadruple ion trap using IR-active ligands.

A strategy for increasing the efficiency of infrared multiphoton dissociation (IRMPD) in a quadrupole ion trap (QIT) is described. IR-active ligands (IRALs) are incorporated into noncovalent complexes of the type [M2+(analyte) IRAL]+, where M is a transition metal such as copper or cobalt and IRAL is an auxiliary ligand with an IR-active phosphonate functional group. The complexes are formed via self-assembly in solution directly prior to ESI-MS analysis. We demonstrate this new IRMPD approach for the structural characterization of flavonoids. The fragment ions obtained by IRMPD are similar to those obtained by CAD and allow facile isomer differentiation of flavonoids. Fourier transform infrared absorption attenuated total reflectance (FTIR-ATR) and energy-variable CAD experiments indicate that the high IRMPD efficiencies stem from the very large IR absorptivities of the IR-active ligands.

Synthesis and characterization of insulin-transferrin conjugates.

Receptor-mediated endocytosis can be exploited for improving the transcellular delivery of therapeutic proteins. Insulin conjugated to transferrin by forming disulfide bonds has been shown to improve insulin oral bioavailability in diabetic rats. We are developing a combination strategy involving complexation hydrogels as delivery vehicles for insulin-transferrin conjugates. The complexation hydrogels developed in our laboratory have been shown to be promising carriers for oral delivery of proteins and peptides. Integrating the strategies based on the complexation hydrogels and insulin-transferrin conjugates may prove to be a novel approach for oral delivery of insulin and other therapeutic proteins. In this work, electrospray ionization mass spectrometry (ESI-MS) was used to study the modification of insulin during its reaction with transferrin. The stability of the conjugated insulin to enzymatic degradation was also studied. ESI-MS studies confirmed the site-specific modifications of insulin. The transferrin conjugation of insulin was also shown to increase the stability of insulin to enzymatic degradation.

Infrared multiphoton dissociation for enhanced de novo sequence interpretation of N-terminal sulfonated peptides in a quadrupole ion trap.

Infrared multiphoton dissociation (IRMPD) of N-terminal sulfonated peptides improves de novo sequencing capabilities in a quadrupole ion trap mass spectrometer. Not only does IRMPD promote highly efficient dissociation of the N-terminal sulfonated peptides but also the entire series of y ions down to the y(1) fragment may be detected due to alleviation of the low-mass cutoff problem associated with conventional collisional activated dissociation (CAD) methods in a quadrupole ion trap. Commercial de novo sequencing software was applied for the interpretation of CAD and IRMPD MS/MS spectra collected for seven unmodified peptides and the corresponding N-terminal sulfonated species. In most cases, the additional information obtained by N-terminal sulfonation in combination with IRMPD provided significant improvements in sequence identification. The software sequence tag results were combined with a commercial database searching algorithm to interpret sequence information of a tryptic digest on alpha-casein s1. Energy-variable CAD studies confirmed a 30-40% reduction in the critical energies of the N-terminal sulfonated peptides relative to unmodified peptides. This reduction in dissociation energy facilitates IRMPD in a quadrupole ion trap.

Verbal abilities as predictors of retention among adolescents in a therapeutic community.

The relationship between verbal skills and retention among adolescents in substance abuse treatment is understudied. In order to assess verbal predictors of retention, twenty-eight 16-19 year old adolescents in a therapeutic community for substance abuse were evaluated between 30 and 90 days after admission. These adolescents were then followed prospectively for 1 year. Verbal and non-verbal cognitive screens, audio taped narrative responses, and self-reports of socio-emotional function and psychiatric symptoms were completed. Verbal scores were associated with self-restraint and counselor reports of therapeutic engagement and comprehension. General verbal scores predicted attrition, while therapeutic expressiveness (verbal expressiveness in a therapeutic context) predicted retention. Remediation of verbal communication skills may be an overlooked aspect of the therapeutic process in treating adolescent substance abusers.

Crystal structure of the CSL-Notch-Mastermind ternary complex bound to DNA.

Notch signaling mediates communication between cells and is essential for proper embryonic patterning and development. CSL is a DNA binding transcription factor that regulates transcription of Notch target genes by interacting with coregulators. Transcriptional activation requires the displacement of corepressors from CSL by the intracellular portion of the receptor Notch (NotchIC) and the recruitment of the coactivator protein Mastermind to the complex. Here we report the 3.1 A structure of the ternary complex formed by CSL, NotchIC, and Mastermind bound to DNA. As expected, the RAM domain of Notch interacts with the beta trefoil domain of CSL; however, the C-terminal domain of CSL has an unanticipated central role in the interface formed with the Notch ankyrin repeats and Mastermind. Ternary complex formation induces a substantial conformational change within CSL, suggesting a molecular mechanism for the conversion of CSL from a repressor to an activator.

Attention-deficit/hyperactivity disorder and early-onset substance use disorders.

In recent years, there has been an increased recognition of the common comorbidity of attention-deficit/ hyperactivity disorder (ADHD) and substance use disorder (SUD) among adolescents and adults. ADHD can be an important factor in the pathogenesis and maintenance of SUD; moreover, retrospective studies suggest that treating ADHD during childhood may prevent the development of SUD. In addition, treatment of ADHD among adults, and possibly adolescents, with SUD can reduce their risk of relapse. Theoretical mechanisms that may explain the relationship between ADHD and SUD are explored in this paper. Current research and recommended clinical practices related to the diagnosis and treatment of ADHD with SUD in adolescents are discussed as well. More research is needed to definitively assess the effectiveness and safety of medications in this population of youths with ADHD and SUD.

Verbal deficits and disruptive behavior disorders among children of opiate-dependent parents.

In order to explore the association between verbal deficits and disruptive behavior disorders among children of addicted parents, 283 6-17-year-old children and their opiate-dependent parents completed diagnostic interviews and standardized measures of vocabulary. Unexpectedly, racial differences in the scores confounded the exploration of the relationship between cognitive scores and disruptive behavior disorders. An interaction between disruptive behavior disorder and race is explored; among Caucasian youths, low verbal scores are associated with disruptive behavior disorders, but this association was not found among African- and Hispanic-American youths. Further analysis and research are needed to understand the clinical significance of relationships between verbal deficits and disruptive behavior disorders within racially diverse groups.

Crystal structure of the dachshund homology domain of human SKI.

The nuclear protooncoprotein SKI negatively regulates transforming growth factor-beta (TGF-beta) signaling in cell growth and differentiation. It directly interacts with the Smads and, by various mechanisms, represses the transcription of TGF-beta-responsive genes. SKI is a multidomain protein that includes a domain bearing high sequence similarity with the retinal determination protein Dachshund (the Dachshund homology domain, DHD). The SKI-DHD has been implicated in SMAD-2/3, N-CoR, SKIP, and PML-RARalpha binding. The 1.65 A crystal structure of the Dachshund homology domain of human SKI is reported here. The SKI-DHD adopts a mixed alpha/beta structure which includes features found in the forkhead/winged-helix family of DNA binding proteins, although SKI-DHD is not a DNA binding domain. Residues that form a contiguous surface patch on SKI-DHD are conserved within the Ski/Sno family and with Dachshund, suggesting that this domain may mediate intermolecular interactions common to these proteins.