Heidelberg, Germany
Heidelberg, Germany

Time filter

Source Type

News Article | April 26, 2017
Site: www.prweb.com

Allergen Control Group Inc. (ACG) is proud to announce a unique collaboration opportunity with Université Laval’s Institute of Nutrition and Functional Foods (INAF) at the Faculty of Agriculture and Food Science, who have initiated the development of a new platform on Food Risk Analysis and Regulatory Excellence (FRAREP). The platform will build on the expertise available at the Institute and Faculty, to support primary knowledge generation and translation in various areas of food risk analysis. The platform aims to gather available information and generate the missing components, to support food regulatory policy proposals which will be substantiated by a risk analysis approach addressing a gap or an emerging food safety and nutrition issue. The goal of the platform is to translate scientific information underpinning food standards and risk management into “a more usable” form for food regulators and industry alike. The intent is to increase the availability and access of ready-to-use food risk analysis information, which is needed by the food community in addressing regulatory challenges. It also aims to become a focal point for a multi-lingual science-driven source of information for food risk communication, adapted to various audiences. Paul Valder, CEO of the Toronto based Allergen Control Group Inc., states that “as leaders in developing and driving voluntary compliance programs for industry, our interest lies in the area of food allergen and gluten management, which has been selected as the first topic for the application of this platform”. The development of food allergen occurrence data, supported by relevant analytical methods and validated by risk assessment methodologies, and the development of adapted risk management approaches are amongst the areas of focus, with the priority on allergens and gluten sources. Under the leadership of Dr. Samuel Godefroy, a former senior food regulator in Canada with extensive international experience, the platform will host its first Center of Expertise in Food Allergen Research (FoARCE), supported by a partnership with food diagnostics global leader r-Biopharm. r-Biopharm is investing an initial $250,000 USD to support applied research efforts aimed at developing, validating and disseminating allergen and gluten detection methodologies to target research initiatives in food allergen and gluten management. Dr. Godefroy is a Full Professor of Food Risk Analysis at the Department of Food Sciences of Université Laval and a researcher of the Institute. “As a collaborative partner, ACG, owners of the risk based Gluten-Free Certification Program, will support the newly established r-Biopharm Canada/ INAF partnership both domestically and internationally to leverage support and maximize the impact of their research outputs, as we help target a leadership role for food allergen and gluten management,” states Valder. For more information about Allergen Control Group and the Platform, please contact: Jessica Burke, Manager, Compliance & Technical Services Allergen Control Group Inc., owners of the Gluten-Free Certification Program (GFCP) jessica.burke(at)glutenfreecert(dot)com


News Article | October 31, 2016
Site: www.marketwired.com

INDIANAPOLIS, IN--(Marketwired - October 31, 2016) - Apexian Pharmaceutical, Inc., a clinical stage biotechnology company focused on developing novel compounds to treat cancer, today announced the hiring of Richard Messmann, M.D. as Chief Medical Officer (CMO) of the company. "We are extremely pleased to have Rich become our CMO at Apexian Pharmaceuticals," said Steve Carchedi, CEO for the company. "Rich is a proven clinical scientist with an outstanding track record in developing oncology products in both large pharma and the biotech firms. Importantly, in his previous positions, Rich has successfully guided many oncology medicines through their clinical development with the goal of benefiting cancer patients." Apexian recently announced acceptance by the U.S. Food and Drug Administration of an Investigational New Drug Application to evaluate the tolerability and anti-tumor effects of APX3330, a small molecule in development to treat cancer. The Phase 1 study will provide important information on the safety of APX3330 in patients with advanced solid tumors and lay the groundwork for subsequent studies. "I am very excited to join the Apexian team as its Chief Medical Officer," said Dr. Messmann. "This is a critically important time for the company in that we are moving into cancer clinical trials with a novel drug backed by an exemplary scientific team led by Dr. Mark R. Kelley at Indiana University School of Medicine. In addition to APX3330, the company has an impressive pipeline of new compounds that we hope will benefit cancer patients in need of safe and effective therapies." Prior to joining Apexian, Rich was CMO of ProNAi Therapeutics and founder of A.B. Biopharm Consulting Group. Rich has over 25 years of experience in cancer drug development including positions at Eli Lilly & Co., Endocyte, Inc., Great Lakes Cancer Institute at Michigan State University and the National Cancer Institute in Bethesda, MD, where he completed a fellowship in medical oncology before becoming a director at the NCI Developmental Therapeutics Program. Apexian Pharmaceuticals, Inc. is a clinical-stage biotechnology company focused on developing novel compounds to treat cancer targeting the multiple functions of the APE1 protein. The lead drug candidate, APX3330, initially targets late stage cancer refractory to existing treatments and is expected to begin clinical studies in 2016. To learn more about Apexian, please visit the Company's website at www.Apexianpharma.com


Schwaerzer G.K.,Free University of Berlin | Hiepen C.,Free University of Berlin | Schrewe H.,Charité - Medical University of Berlin | Schrewe H.,Max Planck Institute for Molecular Genetics | And 5 more authors.
Journal of Bone and Mineral Research | Year: 2012

Growth and differentiation factor 5 (GDF5), a member of the bone morphogenetic protein (BMP) family, is essential for cartilage, bone, and joint formation. Antagonists such as noggin counteract BMP signaling by covering the ligand's BMP type I (BMPRI) and type II (BMPRII, ActRII, ActRIIB) interaction sites. The mutation GDF5-S94N is located within the BMPRII interaction site, the so-called knuckle epitope, and was identified in patients suffering from multiple synostoses syndrome (SYNS). SYNS is characterized by progressive symphalangism, carpal/tarsal fusions, deafness and mild facial dysmorphism. Here we present a novel molecular mechanism of a GDF5 mutation affecting chondrogenesis and osteogenesis. GDF5-S94N exhibits impaired binding to BMPRII causing alleviated Smad and non-Smad signaling and reduced chondrogenic differentiation of ATDC5 cells. Surprisingly, chondrogenesis in mouse micromass cultures was strongly enhanced by GDF5-S94N. By using quantitative techniques (SPR, reporter gene assay, ALP assay, qPCR), we uncovered that this gain of function is caused by strongly reduced affinity of GDF5-S94N to the BMP/GDF antagonist noggin and the consequential lack of noggin inhibition. Thus, since noggin is upregulated during chondrogenic differentiation, GDF5-S94N exceeds the GDF5 action, which results in the phenotypic outcome of SYNS. The detailed molecular characterization of GDF5-S94N as a noggin-resistant growth factor illustrates the potential of GDF5 mutants in applications with defined therapeutical needs. © 2012 American Society for Bone and Mineral Research.


— Age Related Macular Degeneration - Companies Involved in Therapeutics Development are 3SBio Inc, AC Immune SA, Achillion Pharmaceuticals Inc, Aciont Inc, Acucela Inc, AdAlta Ltd, Adverum Biotechnologies Inc, Aerie Pharmaceuticals Inc, Aerpio Therapeutics Inc, Alimera Sciences Inc, Alkeus Pharmaceuticals Inc, Allergan Plc, Allinky Biopharma, Alteogen Inc, Amakem NV, Amarna Therapeutics BV, Ampio Pharmaceuticals Inc, Amyndas Pharmaceuticals LLC, ANP Technologies Inc, Apellis Pharmaceuticals Inc, Apexigen Inc, Applied Genetic Technologies Corp, Astellas Pharma Inc, Benitec Biopharma Ltd, Biokine Therapeutics Ltd, BioMAS Ltd, Biomics Biotechnologies Co Ltd, Biophytis SAS, BLR Bio LLC, Boehringer Ingelheim GmbH, Caladrius Biosciences Inc, Catalyst Biosciences Inc, Cell Cure Neurosciences Ltd, Cell Medica Ltd, Charlesson LLC, Chong Kun Dang Pharmaceutical Corp, Cipla Ltd, Clanotech AB, Clearside BioMedical Inc, Coherus BioSciences Inc, Colby Pharmaceutical Company, Crinetics Pharmaceuticals Inc, Critical Pharmaceuticals Ltd, Daiichi Sankyo Company Ltd, Diffusion Pharmaceuticals Inc, Dong-A Socio Holdings Co Ltd, Eleven Biotherapeutics Inc., Elsalys Biotech SAS, Exonate Ltd, F. Hoffmann-La Roche Ltd, FirstString Research Inc, Foamix Pharmaceuticals Ltd, Formycon AG, Gene Techno Science Co Ltd, Genentech Inc, GenSight Biologics SA, GlaxoSmithKline Plc, Graybug Vision Inc, Grupo Ferrer Internacional SA, HanAll Biopharma Co Ltd, Huabo Biopharm Co Ltd, iCo Therapeutics Inc., Icon Bioscience Inc, Iconic Therapeutics Inc, Inception Sciences Inc, Innovent Biologics Inc, Intellect Neurosciences Inc, International Stem Cell Corp, Ionis Pharmaceuticals Inc, Jeil Pharmaceutical Co Ltd, Johnson & Johnson, Kala Pharmaceuticals Inc, Kodiak Sciences Inc, Lead Discovery Center GmbH, LeadArtis SL, M's Science Corp, Mabion SA, MacuCLEAR Inc, MeiraGTx Ltd, Mitotech SA, Mitsubishi Tanabe Pharma Corp, Mor Research Application Ltd, Navigen Pharmaceuticals Inc, Navya Biologicals Pvt Ltd, Neovacs SA, Neumedicines Inc, Neuroptis Biotech, Novartis AG, NovelMed Therapeutics Inc, Ocular Therapeutix Inc, Ohr Pharmaceutical Inc, Omeros Corp, Ophthotech Corp, Oxford BioMedica Plc, PanOptica Inc, Pfenex Inc, Pfizer Inc, Precision Ocular Ltd, Promedior Inc, pSivida Corp, QLT Inc, Ra Pharmaceuticals Inc, Regeneron Pharmaceuticals Inc, RegenxBio Inc, Retrotope Inc, Ribomic Inc, RXi Pharmaceuticals Corp, Samumed LLC, SanBio Inc, Santen Pharmaceutical Co Ltd, SciFluor Life Sciences LLC, Stealth BioTherapeutics Inc, Sucampo Pharmaceuticals Inc, Sumitomo Dainippon Pharma Co Ltd, Sun Pharma Advanced Research Company Ltd, TRACON Pharmaceuticals Inc, TWi Pharmaceuticals Inc, Tyrogenex Inc, Wellstat Ophthalmics Corp and Xbrane Biopharma AB. Age related macular degeneration is the most common reason for vision loss in people aged above 50. It results in depreciation of the macula that may lead to distorted or blurry central vision. The predisposing factors involved are age, smoking, sunlight, heredity etc. Symptoms include development of blind spot and hazy vision. The condition may be treated by photodynamic therapy, radiation therapy and medication such as anti-angiogenic drugs. The Age Related Macular Degeneration (Ophthalmology) pipeline guide also reviews of key players involved in therapeutic development for Age Related Macular Degeneration and features dormant and discontinued projects. The guide covers therapeutics under Development by Companies /Universities /Institutes, the molecules developed by Companies in Pre-Registration, Phase III, Phase II, Phase I, Preclinical and Discovery stages are 1, 8, 33, 11, 109 and 31 respectively. Similarly, the Universities portfolio in Phase II, Preclinical and Discovery stages comprises 1, 15 and 2 molecules, respectively. Age Related Macular Degeneration (Ophthalmology) pipeline guide helps in identifying and tracking emerging players in the market and their portfolios, enhances decision making capabilities and helps to create effective counter strategies to gain competitive advantage. The guide is built using data and information sourced from Global Markets Direct’s proprietary databases, company/university websites, clinical trial registries, conferences, SEC filings, investor presentations and featured press releases from company/university sites and industry-specific third party sources. Additionally, various dynamic tracking processes ensure that the most recent developments are captured on a real time basis. Inquire more about this research at http://www.reportsnreports.com/contacts/inquirybeforebuy.aspx?name=786897 Note:Certain content / sections in the pipeline guide may be removed or altered based on the availability and relevance of data. • The pipeline guide provides a snapshot of the global therapeutic landscape of Age Related Macular Degeneration (Ophthalmology). • The pipeline guide reviews pipeline therapeutics for Age Related Macular Degeneration (Ophthalmology) by companies and universities/research institutes based on information derived from company and industry-specific sources. • The pipeline guide covers pipeline products based on several stages of development ranging from pre-registration till discovery and undisclosed stages. • The pipeline guide features descriptive drug profiles for the pipeline products which comprise, product description, descriptive licensing and collaboration details, R&D brief, MoA & other developmental activities. • The pipeline guide reviews key companies involved in Age Related Macular Degeneration (Ophthalmology) therapeutics and enlists all their major and minor projects. • The pipeline guide evaluates Age Related Macular Degeneration (Ophthalmology) therapeutics based on mechanism of action (MoA), drug target, route of administration (RoA) and molecule type. • The pipeline guide encapsulates all the dormant and discontinued pipeline projects. • The pipeline guide reviews latest news related to pipeline therapeutics for Age Related Macular Degeneration (Ophthalmology) • Procure strategically important competitor information, analysis, and insights to formulate effective R&D strategies. • Recognize emerging players with potentially strong product portfolio and create effective counter-strategies to gain competitive advantage. • Find and recognize significant and varied types of therapeutics under development for Age Related Macular Degeneration (Ophthalmology). • Classify potential new clients or partners in the target demographic. • Develop tactical initiatives by understanding the focus areas of leading companies. • Plan mergers and acquisitions meritoriously by identifying key players and it’s most promising pipeline therapeutics. • Formulate corrective measures for pipeline projects by understanding Age Related Macular Degeneration (Ophthalmology) pipeline depth and focus of Indication therapeutics. • Develop and design in-licensing and out-licensing strategies by identifying prospective partners with the most attractive projects to enhance and expand business potential and scope. • Adjust the therapeutic portfolio by recognizing discontinued projects and understand from the know-how what drove them from pipeline. For more information, please visit http://www.reportsnreports.com/


Kasten P.,University of Heidelberg | Kasten P.,TU Dresden | Beyen I.,University of Heidelberg | Bormann D.,Institute of Materials Science | And 3 more authors.
Biomaterials | Year: 2010

The osteoinductivity of human growth-and-differentiation factor-5 (GDF-5) is well established, but a reduced amount of ectopic bone is formed compared to other members of the bone morphogenetic protein (BMP) family like BMP-2. We hypothesized that swap of two BMP-receptor-interacting residues of GDF-5 to amino acids present in BMP-2 (methionine to valine at the sites 453 and 456) may improve the bone formation capacity of the mutant GDF-5. Heterotopic bone formation of a mutant GDF-5 coated β-TCP carrier was compared to carriers coated with similar amounts (10 μg) of GDF-5 and BMP-2 in SCID mice. Four week explants revealed 6-fold higher ALP activity in the mutant GDF-5 versus the wild type GDF-5 group (p < 0.0001) and 1.4-fold higher levels compared to BMP-2 (p < 0.006). Bone area in histology was significantly higher in mutant GDF-5 versus all other groups at 4 weeks; however, at 8 weeks BMP-2 reached a similar neo-bone formation like mutant GDF-5. Micro-CT evaluation confirmed higher values in the mutant GDF-5 and BMP-2 groups compared to wild type GDF-5. In conclusion, the mutant GDF-5 showed superior bone formation capacity than GDF-5, and a faster induction at similar final outcome as BMP-2. Mutant GDF-5 thus represents a promising new GDF-5 variant for bone regeneration possibly acting via an increased binding affinity to the BMP-type I receptor. © 2010 Elsevier Ltd. All rights reserved.


Kleinschmidt K.,University of Heidelberg | Kleinschmidt K.,Biologics | Ploeger F.,Biopharm GmbH | Nickel J.,University of Würzburg | And 3 more authors.
Biomaterials | Year: 2013

Non healing bone defects remain a worldwide health problem and still only few osteoinductive growth factors are available for clinical use in bone regeneration. By introducing BMP-2 residues into growth and differentiation factor (GDF)-5 we recently produced a mutant GDF-5 protein BB-1 which enhanced heterotopic bone formation in mice. Designed to combine positive features of GDF-5 and BMP-2, we suspected that this new growth factor variant may improve long bone healing compared to the parent molecules and intended to unravel functional mechanisms behind its action. BB-1 acquired an increased binding affinity to the BMP-IA receptor, mediated enhanced osteogenic induction of human mesenchymal stem cells versus GDF-5 and higher VEGF secretion than BMP-2 invitro. Rabbit radius defects treated with a BB-1-coated collagen carrier healed earlier and with increased bone volume compared to BMP-2 and GDF-5 according to invivo micro-CT follow-up. While BMP-2 callus often remained spongy, BB-1 supported earlier corticalis and marrow cavity formation, showing no pseudojoint persistence like with GDF-5. Thus, by combining positive angiogenic and osteogenic features of GDF-5 and BMP-2, only BB-1 restored a natural bone architecture within 12 weeks, rendering this promising growth factor variant especially promising for long bone regeneration. © 2013 Elsevier Ltd.


PubMed | Target GmbH, German Cancer Research Center, Biopharm GmbH, Copenhagen University and University of Heidelberg
Type: Journal Article | Journal: International journal of cancer | Year: 2016

A small percentage of healthy donors identified in the Western population carry antibodies in their peripheral blood which convey cytotoxic activity against certain human melanoma and neuroblastoma cell lines. We measured the cytotoxic activity of sera and plasmas from healthy donors on the human neuroblastoma cell line Kelly and various melanoma cell lines. Antibodies of IgM isotype, presumably belonging to the class of naturally occurring antibodies, exerted cytotoxic activity in a complement-dependent fashion. Apart from complement-dependent tumor cell lysis, we observed C3 opsonization in all tumor cell lines upon treatment with cytotoxic plasmas. Cell lines tested primarily expressed membrane complement regulatory proteins (mCRP) CD46, CD55 and CD59 to various extents. Blocking of mCRPs by monoclonal antibodies enhanced cell lysis and opsonization, though some melanoma cells remained resistant to complement attack. Epitopes recognized by cytotoxic antibodies were represented by gangliosides such as GD2 and GD3, as evidenced by cellular sialidase pretreatment and enhanced expression of distinct gangliosides. It remains to be clarified why only a small fraction of healthy persons carry these antitumor cytotoxic antibodies.


News Article | December 2, 2015
Site: www.nature.com

No statistical methods were used to predetermine sample size. The Ptet-lacY region of the pZE12 Ptet-lacY plasmid43 was amplified with upstream and downstream primers including the digestion sites XhoI and BamHI, respectively, using the primers Ptet-F and lacY-R (see primers below). The resulting DNA fragment was used to replace the corresponding region of Ptet-gfp in the plasmid pZA31-gfp44, yielding the plasmid pZA31 Ptet-lacY. This plasmid was transformed into the titratable PtsG strain NQ1243 to yield NQ1312. The same procedure was employed to generate the lacYA177V mutant (that is, C531T), but fusion PCR was used to introduce a point mutation Val177 into the lacY sequence33. For this, two overlapping parts of the Ptet-lacY region were PCR amplified with the primers ptet-F, lacYfusion-R and lacYfusion-F, lacY-R (see primers below), in which the point substitution C531T leading to the Val177 mutation from ref. 3 was included in the primers lacYfusion-F and lacYfusion-R. These two overlapping DNA fragments were fused together by PCR using primers ptet-F and lacY-R. The resulting Ptet-lacY fragment that carries the desired mutation was inserted into pZA31, yielding pZA31-lacYA177V. The resulting plasmids were transformed into the titratable PtsG strain NQ1243 to yield NQ1313. The ΔflhD deletion allele in strain JW1881-1 (E. coli Genetic Stock Center, Yale University), in which a kanamycin-resistance gene is substituted for the flhD gene, was transferred to the titratable PtsG strain NQ1243 after deletion of kanamycin resistance by phage P1 vir-mediated transduction. Similarly, the ΔfliA allele from strain JW1907 (KEIO collection45), in which a kanamycin-resistance gene is substituted for the fliA gene, was transferred to the titratable PtsG strain NQ1243 after deletion of kanamycin resistance by phage P1 vir-mediated transduction. The following primers for producing the new genetic constructs were used. ptet-F, 5′-ACACTCGAGTCCCTATCAGTGATAGAGATTG-3′, was used for forward amplification of the Ptet sequence and included an XhoI digestion site for construction of pZE1 Ptetstab-lacZ, pZA31-lacY, pZA31-lacYA177V. lacY-R, 5′-TGTGGATCCTTAAGCGACTTCATTCACCTG-3′, was used for reverse amplification of lacY, lacYA177V and included a BamHI digestion site for construction of pZA31-lacY, pZA31-lacYA177V. lacYfusion-F, 5′-CTCTGGCTGTGTACTCATCCTCGCCGTTTTACTCTTTTTCGCCAAAACGG-3′, was used for forward amplification of a fragment of lacY together with the reverse primer lacY-R. This DNA fragment was later used for fusion PCR to construct pZA31-lacYA177V. lacYfusion-R, 5′-CCGTTTTGGCGAAAAAGAGTAAAACGGCGAGGATGAGTACACAGCCAGAG-3′, was used for reverse amplification of a fragment of Ptet-lacY together with the forward primer ptet-F. This DNA fragment later was used for fusion PCR to construct pZA31-lacYA177V. Our growth media were based on the MOPS-buffered minimal medium used previously46 with slight modifications. The base medium contains 40 mM MOPS and 4 mM tricine (adjusted to pH 7.4 with KOH), 0.1 M NaCl, 10 mM NH Cl, 1.32 mM KH PO , 0.523 mM MgCl , 0.276 mM Na SO , 0.1 mM FeSO and the trace micronutrients described previously47. For 15N-labelled media, 15NH Cl was used in place of 14NH Cl. The concentrations of the carbon sources and various supplements used are indicated in the relevant tables. Batch culture growth has been described in detail previously27. To measure CO production from the bacterial growth, cells were grown in a Multifors bioreactor (Infors HT). Medium (400 ml) was used in a 750-ml vessel, which has an inlet for compressed air and out outlet for the exhaust gas. The vessel is otherwise closed except during brief period of sample collection. Samples of the cell culture (for reading A , assaying lactose and acetate, etc) can be taken by using a syringe connected to the vessel. The air flow rate to the inlet was controlled by a mass flow controller (Cole-Parmer, 32907-67) and maintained at 400 ml min−1. The outlet was connected to a BlueInOne Cell sensor unit (BlueSens) for measuring CO concentration. The stir rate in the growth vessel was set as 800 r.p.m. and temperature was maintained at 37 °C. Samples (100 μl) were taken for at least eight different times during exponential growth (typically at A between 0.1 and 0.6) and immediately frozen. Before the assay, samples were thawed in water and immediately centrifuged at maximum speed (13,200g) for 2.5 min. Supernatant (7 μl) was used to measure glucose concentrations using the Glucose Assay Kit (GAHK-20, Sigma-Aldrich). The slope of the plot of glucose concentrations versus A for all replicates (multiplied with the measured growth rate) was used to determine the glucose uptake rate. To assay lactose, ~10 μl of the collected supernatant was first digested by β-galactosidase (Sigma-Aldrich) in Z-buffer at 37 °C for 20 min. The released glucose was then assayed enzymatically by the kit commercially available (Glucose Assay Kit, GAHK20; Sigma-Aldrich). As a control, the sample was treated in the same way without β-galactosidase. Little glucose was detected in the control. Samples (200 μl) were taken for at least three different times during exponential growth (typically at A between 0.1 and 0.6) and immediately frozen. Before the assay, samples were thawed in water and immediately centrifuged at maximum speed (13,200g) for 2.5 min. Supernatant (100 μl) were used to measure acetate concentrations using the Acetate Assay kit (10148261035, R-Biopharm). The slope of the plot of acetate concentrations versus A for all replicates (multiplied with the measured growth rate) was used to determine the acetate excretion rate. The assay was performed following a similar protocol as detailed in a previous study21. Protein mass spectrometry samples were collected from the four bioreactor cultures, a water bath culture of equation (353) grown on glucose minimal medium, and two 15N-labelled water bath cultures of NCM3722 on lactose minimal medium and NQ381 with 200 μM 3-methylbenzyl alcohol. For each of the cultures, 1.8 ml of cell culture at A  = 0.4–0.5 during the exponential phase was collected by centrifugation. The cell pellet was re-suspended in 0.2 ml water and fast frozen on dry ice. Sample preparation and mass spectrometry methods have been described previously28. The raw mass spectrometry data files generated by the AB SCIEX TripleTOF 5600 system were converted to Mascot generic format (mgf) files, which were submitted to the Mascot database searching engine (Matrix Sciences) against the E. coli SwissProt database to identify proteins. The following parameters were used in the Mascot searches: maximum of two missed trypsin cleavage, fixed carbamidomethyl modification, variable oxidation modification, peptide tolerance ±0.1 daltons (Da), MS/MS tolerance ±0.1 Da, and 1+, 2+ and 3+ peptide charge. All peptides with scores less than the identity threshold (P = 0.05) were discarded. The raw mass spectrometry data files were converted to the .mzML and .mgf formats using conversion tools provided by AB Sciex. The .mgf files were used to identify sequencing events against the Mascot database. Finally, results of the Mascot search were submitted with .mzML files to our in-house quantification software48. In brief, intensity is collected for each peptide over a box in retention time and m/z space that encloses the envelope for the light and heavy peaks. The data are collapsed in the retention time dimension and the light and heavy peaks are fit to a multinomial distribution (a function of the chemical formula of each peptide) using a least squares Fourier transform convolution routine49, which yields the relative intensity of the light and heavy species. The ratio of the non-labelled to labelled peaks was obtained for each peptide in each sample. The relative protein quantification data for each protein in each sample mixture was then obtained as a ratio by taking the median of the ratios of its peptides. No ratio (that is, no data) was obtained if there was only one peptide for the protein. The uncertainty for each ratio was defined as the two quartiles associated with the median. To filter out data with poor quality, the ratio was removed for the protein in that sample if at least one of its quartiles lay outside of 50% range of its median. Furthermore, ratios were removed for a protein in all the sample mixtures in a growth limitation if at least one of the ratios has one of its quartiles lying outside of the 100% range of the median. The spectral counting data used for absolute quantitation were extracted from the Mascot search results. For our 15N and 14N mixture samples, only the 14N spectra were counted. The absolute abundance of a protein was calculated by dividing the total number of 14N spectra of all peptides for that protein by the total number of 14N spectra in the sample. For the condition of the LacZ overexpression strain (NQ1389) grown on glucose medium with zero chlorotetracycline level (see source data file of Fig. 2), 15N sample was prepared, that is, NQ1389 grown on glucose minimal medium with 15NH Cl. The sample was mixed with a known amount of purified LacZ protein (Roche Diagnostics, 10745731001), the purity of which was verified both on a SDS–PAGE gel (where a single band was observed) and by checking the spectral counts of 14N peptides in the sample (where ~99% of the 14N peptides are LacZ peptides). With the highly accurate relative protein abundance between the purified 14N LacZ and the 15N LacZ in the sample, the proteome fraction of LacZ in the sample was determined to be 3.3% ± 0.3%. The average Miller Unit (MU) for the same condition was ~20,550 (see source data file of Fig. 2), leading to a converting factor of 1.6% of proteome fraction for 10,000 MU. Biological replicates show the following typical uncertainties in measured quantities: growth rate, ~5%; acetate excretion rates, ~15%; CO evolution rate, ~5%. The parameters and their associated standard errors for linear relations were obtained by carrying out linear regression. Following our approach, multiple measurements over wide ranges of conditions from robust data sets revealing underlying relations between variables. The uncertainties are reported in Extended Data Tables 2 and 3, and throughout the text.

Loading Biopharm GmbH collaborators
Loading Biopharm GmbH collaborators