FMP Publications

Our publications are recorded in a searchable database since 2010, updates will be added regularly.

Year:  
All :: 2010, 2011, 2012, 2013, ... , 2017
Author:  
All :: (, A, B, C, D, E, F, G, H, I, J, K, L, M, N, O, P, Q, R, S, T, U, V, W, X, Y, Z 
Preferences: 
References per page: Show keywords Show abstracts
References
Dynamic nuclear polarization of deuterated proteins
Akbey, Ü., Franks, W. T., Linden, A., Lange, S., Griffin(*), R. G., van Rossum, B. J.; Oschkinat, H.
Angew Chem Int Ed Engl, 49:7803-7806
(2010)

Tags: Protein Structure (Oschkinat)

New function for an old enzyme: NEP deficient mice develop late-onset obesity
Becker, M., Siems, W. E., Kluge(*), R., Gembardt(*), F., Schultheiss(*), H. P., Schirner(*), M.; Walther(*), T.
Plos One, 5
(2010)

Tags: Biochemical Neurobiology (Siems)

Abstract: BACKGROUND: According to the World Health Organization (WHO) there is a pandemic of obesity with approximately 300 million people being obese. Typically, human obesity has a polygenetic causation. Neutral endopeptidase (NEP), also known as neprilysin, is considered to be one of the key enzymes in the metabolism of many active peptide hormones. METHODOLOGY/PRINCIPAL FINDINGS: An incidental observation in NEP-deficient mice was a late-onset excessive gain in body weight exclusively from a ubiquitous accumulation of fat tissue. In accord with polygenetic human obesity, mice were characterized by deregulation of lipid metabolism, higher blood glucose levels, with impaired glucose tolerance. The key role of NEP in determining body mass was confirmed by the use of the NEP inhibitor candoxatril in wild-type mice that increased body weight due to increased food intake. This is a peripheral and not a central NEP action on the switch for appetite control, since candoxatril cannot cross the blood-brain barrier. Furthermore, we demonstrated that inhibition of NEP in mice with cachexia delayed rapid body weight loss. Thus, lack in NEP activity, genetically or pharmacologically, leads to a gain in body fat. CONCLUSIONS/SIGNIFICANCE: In the present study, we have identified NEP to be a crucial player in the development of obesity. NEP-deficient mice start to become obese under a normocaloric diet in an age of 6-7 months and thus are an ideal model for the typical human late-onset obesity. Therefore, the described obesity model is an ideal tool for research on development, molecular mechanisms, diagnosis, and therapy of the pandemic obesity.

Bidirectional binding of invariant chain peptides to an MHC class II molecule
Günther, S., Schlundt, A., Sticht, J., Roske(*), Y., Heinemann(*), U., Wiesmüller(*), K. H., Jung(*), G., Falk(*), K., Rötzschke(*), O.; Freund, C.
Proc Natl Acad Sci U S A, 107:22219-22224
(2010)

Tags: Protein Engineering (Freund)

Abstract: T-cell recognition of peptides bound to MHC class II (MHCII) molecules is a central event in cell-mediated adaptive immunity. The current paradigm holds that prebound class II-associated invariant chain peptides (CLIP) and all subsequent antigens maintain a canonical orientation in the MHCII binding groove. Here we provide evidence for MHCII-bound CLIP inversion. NMR spectroscopy demonstrates that the interconversion from the canonical to the inverse alignment is a dynamic process, and X-ray crystallography shows that conserved MHC residues form a hydrogen bond network with the peptide backbone in both orientations. The natural catalyst HLA-DM accelerates peptide reorientation and the exchange of either canonically or inversely bound CLIP against antigenic peptide. Thus, noncanonical MHC-CLIP displays the hallmarks of a structurally and functionally intact antigen-presenting complex.

Effects of ACE2 inhibition in the post-myocardial infarction heart
Kim(*), M. A., Yang(*), D., Kida(*), K., Molotkova(*), N., Yeo(*), S. J., Varki(*), N., Iwata(*), M., Dalton(*), N. D., Peterson(*), K. L., Siems, W. E., Walther(*), T., Cowling(*), R. T., Kjekshus(*), J.; Greenberg(*), B.
Journal of cardiac failure, 16:777-785
(2010)

Tags: Biochemical Neurobiology (Siems)

Abstract: BACKGROUND: There is evidence that angiotensin-converting enzyme 2 (ACE2) is cardioprotective. To assess this in the post-myocardial infarction (MI) heart, we treated adult male Sprague-Dawley rats with either placebo (PL) or C16, a selective ACE2 inhibitor, after permanent coronary artery ligation or sham operation. METHODS AND RESULTS: Coronary artery ligation resulting in MI between 25% to 50% of the left ventricular (LV) circumference caused substantial cardiac remodeling. Daily C16 administration from postoperative days 2 to 28 at a dose that inhibited myocardial ACE2 activity was associated with a significant increase in MI size and reduction in LV % fractional shortening. Treatment with C16 did not significantly affect post-MI increases in LV end-diastolic dimension but did inhibit increases in wall thickness and fibrosis in non-infarcted LV. On postoperative day 7, C16 had no significant effect on the increased level of apoptosis in the infarct and border zones nor did it significantly affect capillary density surrounding the MI. It did, however, significantly reduce the number of c-kit(+) cells in the border region. CONCLUSIONS: These findings support the notion that ACE2 exerts cardioprotective effects by preserving jeopardized cardiomyocytes in the border zone. The reduction in hypertrophy and fibrosis with C16, however, suggests that ACE2 activity has diverse effects on post-MI remodeling.

A new phenotype of nongoitrous and nonautoimmune hyperthyroidism caused by a heterozygous thyrotropin receptor mutation in transmembrane helix 6
Winkler(*), F., Kleinau, G., Tarnow(*), P., Rediger(*), A., Grohmann(*), L., Gaetjens(*), I., Krause, G., L'Allemand(*), D., Grüters(*), A., Krude(*), H.; Biebermann(*), H.
The Journal of clinical endocrinology and metabolism, 95:3605-3610
(2010)

Tags: Structural Bioinformatics and Protein Design (Krause, G.)

Abstract: CONTEXT: Activating mutations in the TSHR gene were found in patients suffering from nonautoimmune hyperthyroidism. In the past, it was assumed that thyroid hyperplasia is due to constitutive activation of the Gs/adenylyl cyclase signaling pathway; however, the physiological role of the Gq/11 pathway in this context remains unclear. OBJECTIVE: In this study, we investigated molecular details of the TSHR in a patient with nonautoimmune and nongoitrous hyperthyroidism. RESULTS: We detected a heterozygous mutation in exon 10 of the TSHR gene leading to an exchange of a cysteine residue for tryptophan at amino acid position 636 in transmembrane helix 6. Functional characterization of the mutant receptor revealed a slight reduction of the cell surface expression and TSH induced cAMP accumulation compared to the wild type. Additional observations included a constitutive activation of the Gs-mediated signaling pathway and a simultaneous nearly complete loss-of-function for the Gq/11 pathway after bovine TSH stimulation. Studies on TSHR models suggest significant changes of important amino acid interactions and the overall helix arrangement caused by mutation C636W. CONCLUSION: We report a patient in whom a TSHR mutation leads to nonautoimmune hyperthyroidism due to a mutation that constitutively activates the Gs signaling pathway but additionally completely inhibits the Gq/11 pathway. The absence of goiter in the patient suggests that the Gq/11 pathway is related to thyroid growth and that different signaling pathways are mediated and regulated by TSH. These functional data could be confirmed by reproducible findings of two siblings with a constitutive activation for both pathways.

Essential molecular determinants for thyroid hormone transport and first structural implications for monocarboxylate transporter 8
Kinne(*), A., Kleinau, G., Hoefig(*), C. S., Grüters(*), A., Köhrle, J., Krause, G.; Schweizer(*), U.
J Biol Chem, 285:28054-28063
(2010)

Tags: Structural Bioinformatics and Protein Design (Krause, G.)

Abstract: Monocarboxylate transporter 8 (MCT8, SLC16A2) is a thyroid hormone (TH) transmembrane transport protein mutated in Allan-Herndon-Dudley syndrome, a severe X-linked psychomotor retardation. The neurological and endocrine phenotypes of patients deficient in MCT8 function underscore the physiological significance of carrier-mediated TH transmembrane transport. MCT8 belongs to the major facilitator superfamily of 12 transmembrane-spanning proteins and mediates energy-independent bidirectional transport of iodothyronines across the plasma membrane. Structural information is lacking for all TH transmembrane transporters. To gain insight into structure-function relations in TH transport, we chose human MCT8 as a paradigm. We systematically performed conventional and liquid chromatography-tandem mass spectrometry-based uptake measurements into MCT8-transfected cells using a large number of compounds structurally related to iodothyronines. We found that human MCT8 is specific for L-iodothyronines and requires at least one iodine atom per aromatic ring. Neither thyronamines, decarboxylated metabolites of iodothyronines, nor triiodothyroacetic acid and tetraiodothyroacetic acid, TH derivatives lacking both chiral center and amino group, are substrates for MCT8. The polyphenolic flavonoids naringenin and F21388, potent competitors for TH binding at transthyretin, did not inhibit T(3) transport, suggesting that MCT8 can discriminate its ligand better than transthyretin. Bioinformatic studies and a first molecular homology model of MCT8 suggested amino acids potentially involved in substrate interaction. Indeed, alanine mutation of either Arg(445) (helix 8) or Asp(498) (helix 10) abrogated T(3) transport activity of MCT8, supporting their predicted role in substrate recognition. The MCT8 model allows us to rationalize potential interactions of amino acids including those mutated in patients with Allan-Herndon-Dudley syndrome.

Signaling-sensitive amino acids surround the allosteric ligand binding site of the thyrotropin receptor
Kleinau, G., Haas, A. K., Neumann(*), S., Worth, C. L., Hoyer, I., Furkert, J., Rutz, C., Gershengorn(*), M. C., Schülein, R.; Krause, G.
Faseb j, 24:2347-2354
(2010)

Tags: Structural Bioinformatics and Protein Design (Krause, G.), Protein Trafficking (Schülein)

Abstract: The thyrotropin receptor [thyroid-stimulating hormone receptor (TSHR)], a G-protein-coupled receptor (GPCR), is endogenously activated by thyrotropin, which binds to the extracellular region of the receptor. We previously identified a low-molecular-weight (LMW) agonist of the TSHR and predicted its allosteric binding pocket within the receptor's transmembrane domain. Because binding of the LMW agonist probably disrupts interactions or leads to formation of new interactions among amino acid residues surrounding the pocket, we tested whether mutation of residues at these positions would lead to constitutive signaling activity. Guided by molecular modeling, we performed site-directed mutagenesis of 24 amino acids in this spatial region, followed by functional characterization of the mutant receptors in terms of expression and signaling, measured as cAMP accumulation. We found that mutations V421I, Y466A, T501A, L587V, M637C, M637W, S641A, Y643F, L645V, and Y667A located in several helices exhibit constitutive activity. Of note is mutation M637W at position 6.48 in transmembrane helix 6, which has a significant effect on the interaction of the receptor with the LMW agonist. In summary, we found that a high proportion of residues in several helices surrounding the allosteric binding site of LMW ligands in the TSHR when mutated lead to constitutively active receptors. Our findings of signaling-sensitive residues in this region of the transmembrane bundle may be of general importance as this domain appears to be evolutionarily retained among GPCRs.

Principles and Determinants of G-Protein Coupling by the Rhodopsin-Like Thyrotropin Receptor
Kleinau, G., Jaeschke(*), H., Worth, C. L., Mueller(*), S., Gonzalez(*), J., Paschke(*), R.; Krause, G.
Plos One, 5
(2010)

Tags: Structural Bioinformatics and Protein Design (Krause, G.)

Abstract: In this study we wanted to gain insights into selectivity mechanisms between G-protein-coupled receptors (GPCR) and different subtypes of G-proteins. The thyrotropin receptor (TSHR) binds G-proteins promiscuously and activates both Gs (cAMP) and Gq (IP). Our goal was to dissect selectivity patterns for both pathways in the intracellular region of this receptor. We were particularly interested in the participation of poorly investigated receptor parts. We systematically investigated the amino acids of intracellular loop (ICL) 1 and helix 8 using site-directed mutagenesis alongside characterization of cAMP and IP accumulation. This approach was guided by a homology model of activated TSHR in complex with heterotrimeric Gq, using the X-ray structure of opsin with a bound G-protein peptide as a structural template. We provide evidence that ICL1 is significantly involved in G-protein activation and our model suggests potential interactions with subunits G alpha as well as G beta gamma. Several amino acid substitutions impaired both IP and cAMP accumulation. Moreover, we found a few residues in ICL1 (L440, T441, H443) and helix 8 (R687) that are sensitive for Gq but not for Gs activation. Conversely, not even one residue was found that selectively affects cAMP accumulation only. Together with our previous mutagenesis data on ICL2 and ICL3 we provide here the first systematically completed map of potential interfaces between TSHR and heterotrimeric G-protein. The TSHR/Gq-heterotrimer complex is characterized by more selective interactions than the TSHR/Gs complex. In fact the receptor interface for binding Gs is a subset of that for Gq and we postulate that this may be true for other GPCRs coupling these G-proteins. Our findings support that G-protein coupling and preference is dominated by specific structural features at the intracellular region of the activated GPCR but is completed by additional complementary recognition patterns between receptor and G-protein subtypes.

An unbalanced translocation unmasks a recessive mutation in the follicle-stimulating hormone receptor (FSHR) gene and causes FSH resistance
Kuechler(*), A., Hauffa(*), B. P., Köninger(*), A., Kleinau, G., Albrecht(*), B., Horsthemke(*), B.; Gromoll(*), J.
European journal of human genetics : EJHG, 18:656-661
(2010)

Tags: Structural Bioinformatics and Protein Design (Krause, G.)

Abstract: Follicle-stimulating hormone (FSH) mediated by its receptor (FSHR) is pivotal for normal gametogenesis. Inactivating FSHR mutations are known to cause hypergonadotropic hypogonadism with disturbed follicular maturation in females. So far, only very few recessive point mutations have been described. We report on a 17-year-old female with primary amenorrhea, hypergonadotropic hypogonadism and disturbed folliculogenesis. Chromosome analysis detected a seemingly balanced translocation 46,XX,t(2;8)(p16.3or21;p23.1)mat. FSHR sequence analysis revealed a novel non-synonymous point mutation in exon 10 (c.1760C>A, p.Pro587His), but no wild-type allele. The mutation was also found in the father, but not in the mother. Furthermore, molecular-cytogenetic analyses of the breakpoint region on chromosome 2 showed the translocation to be unbalanced, containing a deletion with one breakpoint within the FSHR gene. The deletion size was narrowed down by array analysis to approximately 163 kb, involving exons 9 and 10 of the FSHR gene. Functional studies of the mutation revealed the complete lack of signal transduction presumably caused by a changed conformational structure of transmembrane helix 6. To our knowledge, this is the first description of a compound heterozygosity of an inactivating FSHR point mutation unmasked by a partial deletion. This coincidence of two rare changes caused clinical signs consistent with FSH resistance.

Disruption of the K+ Channel beta-Subunit KCNE3 Reveals an Important Role in Intestinal and Tracheal Cl- Transport
Preston, P., Wartosch, L., Günzel(*), D., Fromm(*), M., Kongsuphol(*), P., Ousingsawat(*), J., Kunzelmann(*), K., Barhanin(*), J., Warth(*), R.; Jentsch, T. J.
Journal of Biological Chemistry, 285:7165-7175
(2010)

Tags: Physiology and Pathology of Ion Transport (Jentsch

Abstract: The KCNE3 beta-subunit constitutively opens outwardly rectifying KCNQ1 (Kv7.1) K+ channels by abolishing their voltage-dependent gating. The resulting KCNQ1/KCNE3 heteromers display enhanced sensitivity to K+ channel inhibitors like chromanol 293B. KCNE3 was also suggested to modify biophysical properties of several other K+ channels, and a mutation in KCNE3 was proposed to underlie forms of human periodic paralysis. To investigate physiological roles of KCNE3, we now disrupted its gene in mice. kcne3(-/-) mice were viable and fertile and displayed neither periodic paralysis nor other obvious skeletal muscle abnormalities. KCNQ1/KCNE3 heteromers are present in basolateral membranes of intestinal and tracheal epithelial cells where they might facilitate transepithelial Cl- secretion through basolateral recycling of K+ ions and by increasing the electrochemical driving force for apical Cl- exit. Indeed, cAMP-stimulated electrogenic Cl- secretion across tracheal and intestinal epithelia was drastically reduced in kcne3(-/-) mice. Because the abundance and subcellular localization of KCNQ1 was unchanged in kcne3(-/-) mice, the modification of biophysical properties of KCNQ1 by KCNE3 is essential for its role in intestinal and tracheal transport. Further, these results suggest KCNE3 as a potential modifier gene in cystic fibrosis.

Page:  
Previous | 1, 2 | Next
Export as:
BibTeX, XML

Leibniz-Forschungsinstitut für Molekulare Pharmakologie im Forschungsverbund Berlin e.V. (FMP)
Campus Berlin-Buch
Robert-Roessle-Str. 10
13125 Berlin, Germany
+4930 94793 - 100 
+4930 94793 - 109 (Fax)
info(at)fmp-berlin.de

Like many sites, we use cookies to optimize the user's browsing experience. Data Protection OK