Agricultural sustainability does not imply biocenotic sustainability

Abstract. Sustainability is an important quality of the types of agriculture nowadays promoted in central Europe, notably ‘biological agriculture’ and ‘integrated production’. Agronomists, decision-makers and the public generally assume that this agricultural sustainability implies the maintenance of species diversity. However, this assumption often does not hold true. This is shown in a case study of moderately ferti-lized Arrhenatheretum meadows in northern Switzerland. Earlier and more frequent mowing, simultaneous harvesting of all the grasslands in a region, and ecological changes in surrounding arable fields, hedges and other ecosystems often cause a decline in plant and animal species richness, while agricultural yield does not noticeably change. To emphasize the distinction the concept of biocenotic sustainability is proposed for describing the capability of a community to maintain its species composition and structure. For maintaining or attaining biocenotic sustainability. results of modern ecology have to be taken into account, e.g. the theories of island biogeography, minimum viable populations, dispersal, and metapopulations. There is evidence that biocenotic sustainability always implies sustainability of agricultural yield.     

Designing repeat proteins: modular leucine-rich repeat protein libraries based on the mammalian ribonuclease inhibitor family

We present a novel approach to design repeat proteins of the leucine-rich repeat (LRR) family for the generation of libraries of intracellular binding molecules. From an analysis of naturally occurring LRR proteins, we derived the concept to assemble repeat proteins with randomized surface positions from libraries of consensus repeat modules. As a guiding principle, we used the mammalian ribonuclease inhibitor (RI) family, which comprises cytosolic LRR proteins known for their extraordinary affinities to many RNases. By aligning the amino acid sequences of the internal repeats of human, pig, rat, and mouse RI, we derived a first consensus sequence for the characteristic alternating 28 and 29 amino acid residue A-type and B-type repeats. Structural considerations were used to replace all conserved cysteine residues, to define less conserved positions, and to decide where to introduce randomized amino acid residues. The so devised consensus RI repeat library was generated at the DNA level and assembled by stepwise ligation to give libraries of 2-12 repeats. Terminal capping repeats, known to shield the continuous hydrophobic core of the LRR domain from the surrounding solvent, were adapted from human RI. In this way, designed LRR protein libraries of 4-14 LRRs (equivalent to 130-415 amino acid residues) were obtained. The biophysical analysis of randomly chosen library members showed high levels of soluble expression in the Escherichia coli cytosol, monomeric behavior as characterized by gel-filtration, and alpha-helical CD spectra, confirming the success of our design approach.     

The 5.5-year results of MegaOATS – autologous transfer of the posterior femoral condyle: a case-series study

Introduction

Large osteochondral defects of the weight-bearing zones of femoral condyles in young and active patients were treated by autologous transfer of the posterior femoral condyle (large osteochondral autogenous transplantation system (MegaOATS)). The technique presented is a sound and feasible salvage procedure to address large osteochondral defects in weight-bearing zones.

Methods

Thirty-six patients between July 1996 and December 2000 were included. Thirty-three patients (10 females, 23 males) were evaluated by the Lysholm score and X-ray scans. A random sample of 16 individuals underwent magnetic resonance imaging analysis. The average age at the date of surgery was 34.3 (15 to 59) years, and the mean follow up was 66.4 (46 to 98) months. The mean defect size was 6.2 (2 to 10.5) cm², in 27 patients affecting the medial femoral condyle and in six patients affecting the lateral femoral condyle. Trauma or osteochondrosis dissecans were pathogenetic in 82%.

Results

The Lysholm score in all 33 individuals showed a highly significant increase from a preoperative median 49.0 points to a median 86.0 points (P ≤ 0.001). Twenty-seven patients returned to recreational sports. X-ray scans showed a rounding of the osteotomy edge in 24 patients, interpreted as a partial remodelling of the posterior femoral condyle. Preoperative osteoarthritis in 17 individuals was related to significant lower Lysholm scores (P = 0.014), but progression in 17 patients did not significantly influence the score results (P = 0.143). All 16 magnetic resonance imaging examinations showed vital and congruent grafts.

Conclusion

Patients significantly improve in the Lysholm score, in daily-life activity levels and in return to recreational sports. Thirty-one out of 33 patients were comfortable with the results and would undergo the procedure again. The MegaOATS technique is therefore recommended as a salvage procedure for young individuals with large osteochondral defects in the weight-bearing zone of the femoral condyle.     

L1 is sequentially processed by two differently activated metalloproteases and presenilin/gamma-secretase and regulates neural cell adhesion, cell migration, and neurite outgrowth

The immunoglobulin superfamily recognition molecule L1 plays important functional roles in the developing and adult nervous system. Metalloprotease-mediated cleavage of this adhesion molecule has been shown to stimulate cellular migration and neurite outgrowth. We demonstrate here that L1 cleavage is mediated by two distinct members of the disintegrin and metalloprotease family, ADAM10 and ADAM17. This cleavage is differently regulated and leads to the generation of a membrane bound C-terminal fragment, which is further processed through gamma-secretase activity. Pharmacological approaches with two hydroxamate-based inhibitors with different preferences in blocking ADAM10 and ADAM17, as well as loss of function and gain of function studies in murine embryonic fibroblasts, showed that constitutive shedding of L1 is mediated by ADAM10 while phorbol ester stimulation or cholesterol depletion led to ADAM17-mediated L1 cleavage. In contrast, N-methyl-d-aspartate treatment of primary neurons stimulated ADAM10-mediated L1 shedding. Both proteases were able to affect L1-mediated adhesion and haptotactic migration of neuronal cells. In particular, both proteases were involved in L1-dependent neurite outgrowth of cerebellar neurons. Thus, our data identify ADAM10 and ADAM17 as differentially regulated L1 membrane sheddases, both critically affecting the physiological functions of this adhesion protein.     

Cytochrome P450cin (CYP176A1) D241N: Investigating the role of the conserved acid in the active site of cytochrome P450s

P450_cin (CYP176A) is a rare bacterial P450 in that contains an asparagine (Asn242) instead of the conserved threonine that almost all other P450s possess that directs oxygen activation by the heme prosthetic group. However, P450_cin does have the neighbouring, conserved acid (Asp241) that is thought to be involved indirectly in the protonation of the dioxygen and affect the lifetime of the ferric-peroxo species produced during oxygen activation. In this study, the P450_cin D241N mutant has been produced and found to be analogous to the P450_cam D251N mutant. P450_cin catalyses the hydroxylation of cineole to give only (1R)-6β-hydroxycineole and is well coupled (NADPH consumed: product produced). The P450_cin D241N mutant also hydroxylated cineole to produce only (1R)-6β-hydroxycineole, was moderately well coupled (31 ± 3%) but a significant reduction in the rate of the reaction (2% as compared to wild type) was observed. Catalytic oxidation of a variety of substrates by D241N P450_cin were used to examine if typical reactions ascribed to the ferric-peroxo species increased as this intermediate is known to be more persistent in the P450_cam D251N mutant. However, little change was observed in the product profiles of each of these substrates between wild type and mutant enzymes and no products consistent with chemistry of the ferric-peroxo species were observed to increase.     

Improving oral bioavailability of cyclic peptides by N-methylation

The renaissance of peptides in pharmaceutical industry results from their importance in many biological functions. However, low metabolic stability and the lack of oral availability of most peptides is a certain limitation. Whereas metabolic instability may be often overcome by development of small cyclic peptides containing d-amino acids, the very low oral availability of most peptides is a serious limitation for some medicinal applications. The situation is complicated because a twofold optimization – biological activity and oral availability – is required to overcome this problem. Moreover, most simple “rules” for achieving oral availability are not general and are applicable only to limited cases. Many structural modifications for increasing biological activities and metabolic stabilities of cyclic peptides have been described, of which N-alkylation is probably the most common. This mini-review focuses on the effects of N-methylation of cyclic peptides in strategies to optimize bioavailabilities.     

A novel strategy to design binding molecules harnessing the modular nature of repeat proteins

Repeat proteins, such as ankyrin or leucine-rich repeat proteins, are ubiquitous binding molecules, which occur, unlike antibodies, intra- and extracellularly. Their unique modular architecture features repeating structural units (repeats), which stack together to form elongated repeat domains displaying variable and modular target-binding surfaces. Based on this modularity, we developed a novel strategy to generate combinatorial libraries of polypeptides with highly diversified binding specificities. This strategy includes the consensus design of self-compatible repeats displaying variable surface residues and their random assembly into repeat domains. We envision that such repeat protein libraries will be highly valuable sources for novel binding molecules especially suitable for intracellular applications.     

BrdU incorporation reveals DNA replication in non dividing glial cells in the larval abdominal CNS ofDrosophila

Glial cells are of significant importance for central nervous system development and function. In insects, knowledge of the types and development of CNS glia is rather low. This is especially true for postembryonic glial development. Using bromodeoxyuridine incorporation and enhancer trap lines we identified a reproducible spatial and temporal pattern of DNA replicating cells in the abdominal larval CNS (A3-7 neuromeres) ofDrosophila melanogaster. These cells correspond to embryonically established glial cells in that region. Except for a specific subfraction, these cells apparently do not divide during larval life. Similar patterns were found in two otherDrosophila species,D. virilis andD. hydei.