New Firm Creation and Failure: A Matching Approach

We propose that the rate of creation and failure of new firm start-ups can be modelled as a search and matching process, as in labor market matching models. Deriving an "entrepreneurial" Beveridge curve, we show that a successful start-up depends on the efficiency with which entrepreneurial ability is matched with business opportunity, and outline a number of possible applications of this matching approach to assist in formalizing the economics of entrepreneurship.Entrepreneurship, start-ups, labor market matching

Coupling of adenine nucleotide binding and hydrolysis to single- and double-stranded DNA binding determines the topoisomerase activity of reverse gyrase from "Thermotoga maritima"

The aim of this PhD thesis was to functionally characterise the hyperthermophilic topoisomerase IA reverse gyrase from the eubacterium Thermotoga maritima with respect to its unique ATP-dependent positive plasmid supercoiling activity. We set out to apply single molecule FRET to observe conformational changes in the N-terminal helicase-like domain and the C-terminal topoisomerase domain of reverse gyrase during topoisomerase activity. Initially, the protocol for the purification of T. maritima reverse gyrase was improved to yield mg amounts of monomeric enzyme with 95% purity. Requirements for positive plasmid supercoiling by reverse gyrase from T. maritima were defined and optimal reaction conditions were established for the rest of the work. However, no universal nucleotide utilisation pattern for topoisomerase activity of reverse gyrases in general exists and various heterogeneous nucleotides-dependent topoisomerase activities have been reported for reverse gyrases from different organisms. Reverse gyrase from T maritima exhibits no topoisomerase activity in the absence of nucleotides but relaxes plasmids in an ADP- and ADPNP-dependent manner. Reverse gyrase positively supercoils plasmid DNA in the presence of ATP. Surprisingly, hydrolysis of ATPγS efficiently promotes positive supercoiling to a similar extend as ATP hydrolysis. Mutation of the GKT sequence in the Walker A motif of the helicase-like domain renders reverse gyrase inactive for nucleotide hydrolysis and reveals vastly reduced topoisomerase activity. Most interestingly, we observed AMP generation in the presence of short double-stranded DNA substrates and plasmid DNA. During plasmid relaxation, ADP and ADPNP are converted into AMP. The same is true for ATP and ATPγS during positive supercoiling. Possibly, ATP is hydrolysed to AMP by reverse gyrase via intermediately generated ADP and energy may be obtained from ADP cleavage. Further investigating the function of the reverse gyrase domains, we demonstrated that the helicase-like domain is a DNA-stimulated ATPase that harbours all determinants for adenine nucleotide binding and hydrolysis of reverse gyrase. The full-length enzyme shows highly reduced ATPase compared to the isolated helicase-like domain even in the presence of DNA. Consequently, the coupling of DNA binding to ATP hydrolysis is diminished in full-length reverse gyrase and the effect of DNA binding on KM,ATP is much smaller than for the isolated helicase-like domain. Thus, the helicase-like domain of reverse gyrase is a nucleotide-dependent switch. Notably, full-length reverse gyrase from T. maritima binds DNA much more tightly compared to the helicase-like domain and single-stranded DNA binds in a cooperative manner. Artificial 20-, 40- and 60-mer DNA constructs were used to elucidate cooperative binding of reverse gyrase. We show that the binding affinity increases with substrate length. In contrast, the DNA-dependent ATPase activity of T. maritima reverse gyrase decreases with substrate length. Cooperative binding was demonstrated for single-stranded DNA substrates longer than 40 nt and points to possible protein-protein interactions of reverse gyrase that may play a role in the positive supercoiling cycle. In order to observe conformational changes in reverse gyrase with smFRET during the supercoiling cycle, fluorescent labelling is required. However, labelling of reverse gyrase cysteine mutants for smFRET measurements is quite challenging. Reverse gyrase from T. maritima contains eight native cysteines in two putative zinc fingers that have to be intact for positive supercoiling activity. Hence, we improved the conditions accordingly for selective labelling of reverse gyrase cysteine mutants. smFRET measurements indicated that the helicase-like domain might be in a more closed conformation compared to the available crystal structure from A. fulgidus reverse gyrase. Furthermore, the gap between the latch region and the topoisomerase domain is suggested to open during the supercoiling cycle, but the calculated FRET distances correspond to the closed conformation observed in the crystal structure. Conformational changes during the catalytic cycle of reverse gyrase have been predicted for the helicase-like domain and the topoisomerase domain. No conformational changes in the helicase-like domain and the topoisomerase domain are detected with different DNA substrates and adenine nucleotides. Thus, the labelled reverse gyrase mutants may either be inactive under the conditions chosen for smFRET measurements or might have been inactivated by fluorescent labelling. Other positions will be chosen to investigate regions prone to undergo conformational changes during the catalytic cycle without impairing the topoisomerase activity of T. maritima reverse gyrase

Statutory Retirement Age and Lifelong Learning

The employability of an aging population in a world of continuous technical change is top of the political agenda. Due to endogenous human capital depreciation, the effective retirement age is often below statutory retirement age resulting in unemployment among older workers. We analyze this phenomenon in a putty-putty human capital vintage model and focus on education and the speed of human capital depreciation. Introducing a two-stage education system with initial schooling and lifelong learning, not even lifelong learning turns out to be capable of aligning economic and statutory retirement. However, lifelong learning can increase the number of people reaching statutory retirement age and hence reduce the problem of old age unemployment in an aging society.lifelong learning, retirement, unemployment, education system

Economic Retirement Age and Lifelong Learning - a theoretical model with heterogeneous labor, biased technical change and international sourcing

The employability of an aging population in a world of continuous and biased technical change is top of the political agenda. Due to endogenous human capital depreciation the effective retirement age is often below statutory retirement age resulting in permanent non-employability of older workers. We analyze this phenomenon in a putty-putty human capital vintage model and focus on education and the speed of human capital depreciation. Introducing a two-stage education system with initial schooling and lifelong learning, not even lifelong learning turns out to be capable of aligning economic and statutory retirement. However, well designed education programs will keep more workers in highly productive activities at the end of their working life, and hence will substitute for simple social transfers, or for an early switch towards very low paid jobs

Crystal structures of Thermotoga maritima reverse gyrase: inferences for the mechanism of positive DNA supercoiling

Reverse gyrase is an ATP-dependent topoisomerase that is unique to hyperthermophilic archaea and eubacteria. The only reverse gyrase structure determined to date has revealed the arrangement of the N-terminal helicase domain and the C-terminal topoisomerase domain that intimately cooperate to generate the unique function of positive DNA supercoiling. Although the structure has elicited hypotheses as to how supercoiling may be achieved, it lacks structural elements important for supercoiling and the molecular mechanism of positive supercoiling is still not clear. We present five structures of authentic Thermotoga maritima reverse gyrase that reveal a first view of two interacting zinc fingers that are crucial for positive DNA supercoiling. The so-called latch domain, which connects the helicase and the topoisomerase domains is required for their functional cooperation and presents a novel fold. Structural comparison defines mobile regions in parts of the helicase domain, including a helical insert and the latch that are likely important for DNA binding during catalysis. We show that the latch, the helical insert and the zinc fingers contribute to the binding of DNA to reverse gyrase and are uniquely placed within the reverse gyrase structure to bind and guide DNA during strand passage. A possible mechanism for positive supercoiling by reverse gyrases is presente

Link between Organ-specific Antigen Processing by 20S Proteasomes and CD8+ T Cell–mediated Autoimmunity

Adoptive transfer of cross-reactive HSP60-specific CD8+ T cells into immunodeficient mice causes autoimmune intestinal pathology restricted to the small intestine. We wondered whether local immunopathology induced by CD8+ T cells can be explained by tissue-specific differences in proteasome-mediated processing of major histocompatibility complex class I T cell epitopes. Our experiments demonstrate that 20S proteasomes of different organs display a characteristic composition of α and β chain subunits and produce distinct peptide fragments with respect to both quality and quantity. Digests of HSP60 polypeptides by 20S proteasomes show most efficient generation of the pathology related CD8+ T cell epitope in the small intestine. Further, we demonstrate that the organ-specific potential to produce defined T cell epitopes reflects quantities that are relevant for cytotoxic T lymphocyte recognition. We propose tissue-specific antigen processing by 20S proteasomes as a potential mechanism to control organ-specific immune responses

The reverse gyrase helicase-like domain is a nucleotide-dependent switch that is attenuated by the topoisomerase domain

Reverse gyrase is a topoisomerase that introduces positive supercoils into DNA in an ATP-dependent manner. It is unique to hyperthermophilic archaea and eubacteria, and has been proposed to protect their DNA from damage at high temperatures. Cooperation between its N-terminal helicase-like and the C-terminal topoisomerase domain is required for positive supercoiling, but the precise role of the helicase-like domain is currently unknown. Here, the characterization of the isolated helicase-like domain from Thermotoga maritima reverse gyrase is presented. We show that the helicase-like domain contains all determinants for nucleotide binding and ATP hydrolysis. Its intrinsic ATP hydrolysis is significantly stimulated by ssDNA, dsDNA and plasmid DNA. During the nucleotide cycle, the helicase-like domain switches between high- and low-affinity states for dsDNA, while its affinity for ssDNA in the ATP and ADP states is similar. In the context of reverse gyrase, the differences in DNA affinities of the nucleotide states are smaller, and the DNA-stimulated ATPase activity is strongly reduced. This inhibitory effect of the topoisomerase domain decelerates the progression of reverse gyrase through the nucleotide cycle, possibly providing optimal coordination of ATP hydrolysis with the complex reaction of DNA supercoiling

Differential regulation of amidase- and formamidase-mediated ammonia production by the Helicobacter pylori fur repressor.

The production of high levels of ammonia allows the human gastric pathogen Helicobacter pylori to survive the acidic conditions in the human stomach. H. pylori produces ammonia through urease-mediated degradation of urea, but it is also able to convert a range of amide substrates into ammonia via its AmiE amidase and AmiF formamidase enzymes. Here data are provided that demonstrate that the iron-responsive regulatory protein Fur directly and indirectly regulates the activity of the two H. pylori amidases. In contrast to other amidase-positive bacteria, amidase and formamidase enzyme activities were not induced by medium supplementation with their respective substrates, acrylamide and formamide. AmiE protein expression and amidase enzyme activity were iron-repressed in H. pylori 26695 but constitutive in the isogenic fur mutant. This regulation was mediated at the transcriptional level via the binding of Fur to the amiE promoter region. In contrast, formamidase enzyme activity was not iron-repressed but was significantly higher in the fur mutant. This effect was not mediated at the transcriptional level, and Fur did not bind to the amiF promoter region. These roles of Fur in regulation of the H. pylori amidases suggest that the H. pylori Fur regulator may have acquired extra functions to compensate for the absence of other regulatory systems

The Swiss Approach - feasibility of a national low-dose CT lung cancer screening program.

BACKGROUND Lung cancer is the leading cause of cancer-related deaths in Switzerland. Despite this, there is no lung cancer screening program in the country. In the United States, low-dose computed tomography (LDCT) lung cancer screening is partially established and endorsed by guidelines. Moreover, evidence is growing that screening reduces lung cancer-related mortality and this was recently shown in a large European randomized controlled trial. Implementation of a lung cancer screening program, however, is challenging and depends on many country-specific factors. The goal of this article is to outline a potential Swiss lung cancer screening program. FRAMEWORK An exhaustive literature review on international screening models as well as interviews and site visits with international experts were initiated. Furthermore, workshops and interviews with national experts and stakeholders were conducted to share experiences and to establish the basis for a national Swiss lung cancer screening program. SCREENING APPROACH General practitioners, pulmonologists and the media should be part of the recruitment process. Decentralisation of the screening might lead to a higher adherence rate. To reduce stigmatisation, the screening should be integrated in a "lung health check". Standardisation and a common quality level are mandatory. The PLCOm2012 risk calculation model with a threshold of 1.5% risk for developing cancer in the next six years should be used in addition to established inclusion criteria. Biennial screening is preferred. LUNG RADS and NELSON+ are applied as classification models for lung nodules. CONCLUSION Based on data from recent studies, literature research, a health technology assessment, the information gained from this project and a pilot study the Swiss Interest Group for lung cancer screening (CH-LSIG) recommends the timely introduction of a systematic lung cancer screening program in Switzerland. The final decision is for the Swiss Cancer Screening Committee to make

The Swiss Approach - feasibility of a national low-dose CT lung cancer screening program

BACKGROUND Lung cancer is the leading cause of cancer-related deaths in Switzerland. Despite this, there is no lung cancer screening program in the country. In the United States, low-dose computed tomography (LDCT) lung cancer screening is partially established and endorsed by guidelines. Moreover, evidence is growing that screening reduces lung cancer-related mortality and this was recently shown in a large European randomized controlled trial. Implementation of a lung cancer screening program, however, is challenging and depends on many country-specific factors. The goal of this article is to outline a potential Swiss lung cancer screening program. FRAMEWORK An exhaustive literature review on international screening models as well as interviews and site visits with international experts were initiated. Furthermore, workshops and interviews with national experts and stakeholders were conducted to share experiences and to establish the basis for a national Swiss lung cancer screening program. SCREENING APPROACH General practitioners, pulmonologists and the media should be part of the recruitment process. Decentralisation of the screening might lead to a higher adherence rate. To reduce stigmatisation, the screening should be integrated in a "lung health check". Standardisation and a common quality level are mandatory. The PLCOm2012 risk calculation model with a threshold of 1.5% risk for developing cancer in the next six years should be used in addition to established inclusion criteria. Biennial screening is preferred. LUNG RADS and NELSON+ are applied as classification models for lung nodules. CONCLUSION Based on data from recent studies, literature research, a health technology assessment, the information gained from this project and a pilot study the Swiss Interest Group for lung cancer screening (CH-LSIG) recommends the timely introduction of a systematic lung cancer screening program in Switzerland. The final decision is for the Swiss Cancer Screening Committee to make