Franz X Schmid

University of Bayreuth, Bayreuth, Bavaria, Germany

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Publications (261)1139.3 Total impact

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    ABSTRACT: The signal adapter protein c-CrkII from chicken but not from human uses isomerization at Pro238 in the SH3C domain to regulate the activity of the SH3N domain. The different behavior of human and chicken c-CrkII originates from only two differences in sequence, at the positions 239 after Pro238 and 272 in the N-Src loop of SH3C. We analyzed the kinetics of substrate binding to SH3N and an assay for its coupling with Pro238 isomerization in SH3C to identify the molecular path from Pro238 to the substrate binding site of SH3N. The trans → cis isomerization at Pro238 and a relocation of Phe239 re-organize the energetics of a hydrophobic cluster in the N-Src loop of SH3C and reshape this region to optimize its interactions with SH3N. Concomitantly, the backbone becomes strained at Met272. We suggest that, in human c-CrkII, movement at position 239 and strain at position 272 are not tolerated, because the β-branched residues Ile239 and Val272 restrain the backbone mobility and thus destabilize the cis Pro238 form.
    No preview · Article · Oct 2015 · Journal of Molecular Biology
  • Philipp A M Schmidpeter · Franz X Schmid
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    ABSTRACT: Prolyl isomerizations are intrinsically slow processes. They determine the rates of many protein folding reactions and control regulatory events in folded proteins. Prolyl isomerases are able to catalyze these isomerizations, and thus they have the potential to assist protein folding and to modulate protein function. Here we provide examples for how prolyl isomerizations limit protein folding and are accelerated by prolyl isomerases, and how native-state prolyl isomerizations regulate protein functions. The roles of prolines in protein folding and protein function are closely interrelated because they both depend on the coupling between cis/trans isomerization and conformational changes that can involve extended regions of a protein. Copyright © 2015. Published by Elsevier Ltd.
    No preview · Article · Feb 2015 · Journal of Molecular Biology
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    ABSTRACT: Background: Prolyl cis/trans isomerizations have long been known as critical and rate-limiting steps in protein folding. Results: Now it is clear that they are also used as slow conformational switches and molecular timers in the regulation of protein activity. Here we describe several such proline switches and how they are regulated. Conclusions and general significance: Prolyl isomerizations can function as attenuators and provide allosteric systems with a molecular memory. This article is part of a Special Issue entitled Proline-directed Foldases: Cell Signaling Catalysts and Drug Targets.
    No preview · Article · Dec 2014 · Biochimica et Biophysica Acta (BBA) - General Subjects
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    ABSTRACT: Secretion of proteins into the membrane-cell wall space is essential for cell wall biosynthesis and pathogenicity in Gram-positive bacteria. Folding and maturation of many secreted proteins depend on a single extracellular foldase, the PrsA protein. PrsA is a 30 kDa protein, lipid-anchored to the outer leaflet of the cell membrane. The crystal structure of Bacillus subtilis PrsA reveals a central catalytic parvulin-type prolyl isomerase domain, which is inserted into a larger composite NC domain formed by the N- and C-terminal regions. This domain architecture resembles, despite a lack of sequence conservation, both trigger factor, a ribosome-binding bacterial chaperone, and SurA, a periplasmic chaperone in Gram-negative bacteria. Two main structural differences are observed in that the N-terminal arm of PrsA is substantially shortened relative to trigger factor and SurA and in that PrsA is found to dimerize in a unique fashion via its NC domain. Dimerization leads to a large, bowl-shaped crevice, which might be involved in vivo in protecting substrate proteins from aggregation. NMR experiments reveal a direct, dynamic interaction of both the parvulin and the NC domain with secretion propeptides, which have been implicated in substrate targeting to PrsA. Copyright © 2014, The American Society for Biochemistry and Molecular Biology.
    No preview · Article · Dec 2014 · Journal of Biological Chemistry
  • Philipp A M Schmidpeter · Franz X Schmid
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    ABSTRACT: c-CrkII is a central signal adapter protein. A domain opening/closing reaction between its N-terminal and C-terminal SH3 domains (SH3N and SH3C, respectively) controls signal propagation from upstream tyrosine kinases to downstream targets. In chicken but not in human c-CrkII, opening/closing is coupled with cis/trans isomerization at Pro238 in SH3C. Here, we used advanced double-mixing experiments and kinetic simulations to uncover dynamic domain interactions in c-CrkII, to elucidate how they are linked with cis/trans isomerization, and how this regulates substrate binding to SH3N. Pro238 trans → cis isomerization is not a simple on/off switch, but converts chicken c-CrkII from a high-affinity to a low-affinity form. We present a double-box model that describes c-CrkII as an allosteric system consisting of an open, high-affinity R state and a closed, low-affinity T state. Coupling of the T-R transition with an intrinsically slow prolyl isomerization provides c-CrkII with a kinetic memory and possibly functions as a molecular attenuator during signal transduction. Copyright © 2014, The American Society for Biochemistry and Molecular Biology.
    No preview · Article · Dec 2014 · Journal of Biological Chemistry
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    ABSTRACT: Trigger factor is the first molecular chaperone interacting cotranslationally with virtually all nascent polypeptides synthesized by the ribosome in bacteria. Thermal adaptation of chaperone function was investigated in trigger factors from the Antarctic psychrophile Pseudoalteromonas haloplanktis, the mesophile Escherichia coli, and the hyperthermophile Thermotoga maritima. This series covers nearly all temperatures encountered by bacteria. Although structurally homologous, these trigger factors display strikingly distinct properties that are related to the bacterial environmental temperature. The hyperthermophilic trigger factor strongly binds model proteins during their folding and protects them from heat-induced misfolding and aggregation. It decreases the folding rate and counteracts the fast folding rate imposed by high temperature. It also functions as a carrier of partially folded proteins for delivery to downstream chaperones ensuring final maturation. By contrast, the psychrophilic trigger factor displays weak chaperone activities showing that these functions are less important in cold conditions, because protein folding, misfolding, and aggregation are slowed down at low temperature. It efficiently catalyzes prolyl isomerization at low temperature as a result of its increased cellular concentration rather than from an improved activity. Some chaperone properties of the mesophilic trigger factor possibly reflect its function as a cold shock protein in Escherichia coli.
    No preview · Article · Nov 2014 · Environmental Microbiology
  • Johanna R. Koch · Franz X. Schmid
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    ABSTRACT: Mia40 (a mitochondrial import and assembly protein) catalyzes disulfide bond formation in proteins in the mitochondria] intermembrane space. By using Cox17 (a mitochondrial copper-binding protein) as a natural substrate, we discovered that, in the presence of Mia40, the formation of native disulfides is strongly favored. The catalytic mechanism of Mia40 involves a functional interplay between the chaperone site and the catalytic disulfide. Mia40 forms a specific native disulfide in Cox17 much more rapidly than other disulfides, in particular, non-native ones, which originates from the recently described high affinity for hydrophobic regions near target cysteines and the long lifetime of the mixed disulfide. In addition to its thiol oxidase function, Mia40 is active also as a disulfide reductase and isomerase. We found that species with inadvertently formed incorrect disulfides are rebound by Mia40 and reshuffled, revealing a proofreading mechanism that is steered by the conformational folding of the substrate protein.
    No preview · Article · Nov 2014 · Journal of Molecular Biology
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    Franz X. Schmid

    Preview · Article · Nov 2014 · Journal of Molecular Biology
  • Philipp A M Schmidpeter · Franz X Schmid
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    ABSTRACT: The signaling protein CrkII switches between forms with high or low binding affinity. Phosphorylation and native-state prolyl isomerization were both suggested to regulate the transition between these forms. Here we analyzed how phosphorylation at Tyr222 and Tyr252, and the Pro238Ala substitution affect signal transfer of human and chicken CrkII to a downstream target. Human CrkII is regulated by phosphorylation only, chicken CrkII by Pro238 trans→cis isomerization as well as by Tyr222 phosphorylation. Surprisingly, they act in an independent fashion. Apparently, the allosteric transition to a low-activity form can be induced by phosphorylation or prolyl isomerization located at distant sites in CrkII.
    No preview · Article · Oct 2014 · Journal of Molecular Biology
  • Johanna R Koch · Franz X Schmid
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    ABSTRACT: Mia40 catalyzes oxidative protein folding in mitochondria. It contains a unique catalytic CPC dithiol flanked by a hydrophobic groove, and unlike other oxidoreductases, it forms long-lived mixed disulfides with substrates. We show that this distinctive property originates neither from particular properties of mitochondrial substrates nor from the CPC motif of Mia40. The catalytic cysteines of Mia40 display unusually low chemical reactivity, as expressed in conventional pK values and reduction potentials. The stability of the mixed disulfide intermediate is coupled energetically with hydrophobic interactions between Mia40 and the substrate. Based on these properties, we suggest a mechanism for Mia40, where the hydrophobic binding site is employed to select a substrate thiol for forming the initial mixed disulfide. Its long lifetime is used to retain partially folded proteins in the mitochondria and to direct folding towards forming the native disulfide bonds.
    No preview · Article · Jul 2014 · ACS Chemical Biology
  • Philipp A M Schmidpeter · Franz X Schmid
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    ABSTRACT: The cellular CT10 regulator of kinase protein (c-CrkII) transmits signals from oncogenic tyrosine kinases to cellular targets. NMR studies had suggested that in chicken c-CrkII a native-state prolyl cis/trans isomerization is involved in signal propagation. Corresponding evidence for the closely related human c-CrkII was not obtained. Here we analyzed the kinetics of folding and substrate binding of the two homologs and found that cis/trans isomerization of Pro238 determines target binding in chicken but not in human c-CrkII. A reciprocal mutational analysis uncovered residues that determine the isomeric state at Pro238 and transmit it to the binding site for downstream target proteins. Transfer of these key residues to human c-CrkII established a regulatory proline switch in this protein as well. We suggest that Pro238 isomerization extends the life time of the signaling-active state of c-CrkII and thereby functions as a long-term molecular storage device.
    No preview · Article · Feb 2014 · ACS Chemical Biology
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    ABSTRACT: To initiate infection of Escherichia coli, phage fd uses its gene-3-protein (G3P) to bind first to an F pilus and then to the TolA protein at the cell surface. G3P is normally auto-inhibited, because a tight interaction between the two N-terminal domains N1 and N2 buries the TolA binding site. Binding of N2 to the pilus activates G3P by initiating long-range conformational changes that are relayed to the domain interface and to a proline timer. We discovered that the 23-28 loop of the N1 domain is critical for propagating these conformational signals. The analysis of the stability and the folding dynamics of G3P variants with a shortened loop, combined with TolA interaction studies and phage infection experiments reveal how the contact between the N2 domain and the 23-28 loop of N1 is energetically linked with the interdomain region and the proline timer and how it affects phage infectivity. Our results illustrate how conformational transitions and prolyl cis/trans isomerization can be coupled energetically and how conformational signals to and from prolines can be propagated over long distances in proteins.
    No preview · Article · Jan 2014 · Journal of Molecular Biology
  • Johanna R Koch · Franz X Schmid
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    ABSTRACT: Mia40 catalyses the oxidative folding of disulphide-containing proteins in the mitochondria. The folding pathway is directed by the formation of the first mixed disulphide between Mia40 and its substrate. Here, we employ Cox17 to elucidate the molecular determinants of this reaction. Mia40 engages initially in a dynamic non-covalent enzyme-substrate complex that forms and dissociates within milliseconds. Cys36 of Cox17 forms the mixed disulphide in an extremely rapid reaction that is limited by the preceding complex formation with Mia40. Cys36 reacts much faster than the three other cysteines of Cox17, because it neighbours three hydrophobic residues. Mia40 binds preferentially to hydrophobic regions and the dynamic nature of the non-covalent complex allows rapid reorientation for an optimal positioning of the reactive cysteine. Mia40 thus uses the unique proximity between its substrate-binding site and the catalytic disulphide to select a particular cysteine for forming the critical initial mixed disulphide.
    No preview · Article · Jan 2014 · Nature Communications
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    Anne-Juliane Geitner · Edina Varga · Marc Wehmer · Franz Xaver Schmid
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    ABSTRACT: Parvulins are small prolyl isomerases and serve as catalytic domains of folding enzymes. SurA from the periplasm of Escherichia coli consists of an inactive (Par1) and an active (Par2) parvulin domain as well as a chaperone domain. In the absence of the chaperone domain, the folding activity of Par2 is virtually abolished. We created a chimeric protein by inserting the chaperone domain of SlyD, an unrelated folding enzyme from the FKBP family, into a loop of the isolated Par2 domain of SurA. This increased its folding activity 450-fold to a value higher than the activity of SurA, in which Par2 is linked with its natural chaperone domain. In the presence of both the natural and the foreign chaperone domain, the folding activity of Par2 was 1500-fold increased. Related and unrelated chaperone domains thus are similarly efficient in enhancing the folding activity of the prolyl isomerase Par2. A sequence analysis of various chaperone domains suggests that clusters of exposed methionine residues in mobile chain regions might be important for a generic interaction with unfolded protein chains. This binding is highly dynamic to allow frequent transfer of folding protein chains between chaperone and catalytic domains.
    Preview · Article · Jul 2013 · Journal of Molecular Biology
  • Michael Kovermann · Franz X Schmid · Jochen Balbach
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    ABSTRACT: Abstract SlyD is a bacterial two-domain protein that functions as a molecular chaperone, a prolyl cis/trans isomerase, and a nickel binding protein. This review summarizes recent findings about the molecular enzyme mechanism of SlyD. The chaperone function located in one domain of SlyD is involved in twin-arginine translocation and increases the catalytic efficiency of the prolyl cis/trans isomerase domain in protein folding by two orders of magnitude. The C-terminal tail of SlyD binds Ni2+ ions and supplies them for the maturation of [NiFe] hydrogenases. A combined biochemical and biophysical analysis revealed the molecular basis of the delicate interplay of the different domains of SlyD for an optimal function.
    No preview · Article · Mar 2013 · Biological Chemistry
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    ABSTRACT: Infection of Escherichia coli by the filamentous phage fd starts with the binding of the N2 domain of the phage gene-3-protein to an F pilus. This interaction triggers partial unfolding of the gene-3-protein, cis/trans isomerization at Pro213, and domain disassembly, thereby exposing its binding site for the ultimate receptor TolA. The trans proline sets a molecular timer to maintain the binding-active state long enough for the phage to interact with TolA. We elucidated the changes in structure and local stability that lead to partial unfolding and thus to the activation of the gene-3-protein for phage infection. Protein folding and TolA binding experiments were combined with real-time NMR spectroscopy, amide hydrogen exchange measurements and phage infectivity assays. In combination, the results provide a molecular picture of how a local unfolding reaction couples with prolyl isomerization not only to generate the activated state of a protein but also to maintain it for an extended time.
    Full-text · Article · Mar 2013 · Journal of Biological Chemistry
  • Gabriel Zoldak · Anne-Juliane Geitner · Franz X Schmid
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    ABSTRACT: Folding enzymes often use distinct domains for the interaction with a folding protein chain and for the catalysis of intrinsically slow reactions such as prolyl cis/trans isomerization. Here we investigated the refolding reaction of ribonuclease T1 in the presence of the prolyl isomerase SlyD from Escherichia coli to examine how this enzyme catalyzes the folding of molecules with an incorrect trans proline isomer and how it modulates the conformational folding of the molecules with the correct cis proline. The kinetic analysis suggests that prolyl cis-trans isomerization in the SlyD-bound state shows a rate near 100 s-1 and is thus more than 10exp4-fold accelerated, relative to the uncatalyzed reaction. As a consequence of its fast binding and efficient catalysis, SlyD retards the conformational folding of the protein molecules with the correct cis isomer, because it promotes the formation of the species with the incorrect trans isomer. In the presence of 1 microM SlyD or higher, protein molecules with cis and trans prolyl isomers refold with identical rates, because SlyD-catalyzed cis/trans equilibration is faster than conformational folding. The cis or trans state of a particular proline is thus no longer a determinant for the rate of folding.
    No preview · Article · Feb 2013 · Journal of the American Chemical Society
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    ABSTRACT: Human aryl hydrocarbon receptor (AHR) interacting protein (AIP) and AIP like 1 (AIPL1) are cochaperones of Hsp90 which share 49% sequence identity. Both proteins contain an N-terminal FKBP-like prolyl peptidyl isomerase (PPIase) domain followed by a Tetratricopeptide repeat (TPR) domain. In addition, AIPL1 harbors a unique C-terminal proline-rich domain (PRD). Little is known about the functional relevance of the individual domains and how these contribute to the association with Hsp90. In this study, we show that these cochaperones differ from other Hsp90-associated PPIase as their FKBP domains are enzymatically inactive. Furthermore, in contrast to other large PPIases, AIP is inactive as a chaperone. AIPL1, however, exhibits chaperone activity and prevents the aggregation of non-native proteins. The unique proline-rich domain of AIPL1 is important for its chaperone function as its truncation severely affects the ability of AIPL1 to bind non-native proteins. Furthermore, the proline-rich domain decreased the affinity of AIPL1 for Hsp90, implying that this domain acts as a negative regulator of the Hsp90 interaction besides being necessary for efficient binding of AIPL1 to non-native proteins.
    No preview · Article · Feb 2013 · Biochemistry
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    ABSTRACT: In general, β-lactamases of medically important Gram-negative bacteria are Sec-dependently translocated into the periplasm. In contrast, β-lactamases of Mycobacteria spp. (BlaC, BlaS) and the Gram-negative environmental bacteria Stenotrophomonas maltophilia (L2) and Xanthomonas campestris (Bla(XCC-1)) have been reported to be secreted by the twin-arginine translocation (Tat) system. Yersinia enterocolitica carries 2 distinct β-lactamase genes (blaA and blaB) encoding BlaA(Ye) and the AmpC-like β-lactamase BlaB, respectively. By using the software PRED-TAT for prediction and discrimination of Sec from Tat signal peptides, we identified a functional Tat signal sequence for Yersinia BlaA(Ye). The Tat-dependent translocation of BlaA(Ye) could be clearly demonstrated by using a Y. enterocolitica tatC-mutant and cell fractionation. Moreover, we could demonstrate a unique unusual temperature-dependent activity profile of BlaA(Ye) ranging from 15 to 60°C and a high 'melting temperature' (T(M)=44.3°) in comparison to the related Sec-dependent β-lactamase TEM-1 (20-50°C, T(M)=34.9°C). Strikingly, the blaA gene of Y. enterocolitica is present in diverse environmental Yersinia spp. and a blaA homolog gene could be identified in the closely related Photorhabdus asymbiotica (BlaA(Pa); 69% identity to BlaA(Ye)). For BlaA(Pa) of P. asymbiotica, we could also demonstrate Tat-dependent secretion. These results suggest that Yersinia BlaA-related β-lactamases may be the prototype of a large Tat-dependent β-lactamase family, which originated from environmental bacteria.
    No preview · Article · Dec 2012 · International journal of medical microbiology: IJMM
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    ABSTRACT: The (βα)(8)-barrel is among the most ancient, frequent, and versatile enzyme structures. It was proposed that modern (βα)(8)-barrel proteins have evolved from an ancestral (βα)(4)-half-barrel by gene duplication and fusion. We explored whether the mechanism of protein folding has remained conserved during this long-lasting evolutionary process. For this purpose, potential primordial (βα)(8)-barrel proteins were constructed by the duplication of a (βα)(4) element of a modern (βα)(8)-barrel protein, imidazole glycerol phosphate synthase (HisF), followed by the optimization of the initial construct. The symmetric variant Sym1 was less stable than HisF and its crystal structure showed disorder in the contact regions between the half-barrels. The next generation variant Sym2 was more stable than HisF, and the contact regions were well resolved. Remarkably, both artificial (βα)(8)-barrels show the same refolding mechanism as HisF and other modern (βα)(8)-barrel proteins. Early in folding, they all equilibrate rapidly with an off-pathway species. On the productive folding path, they form closely related intermediates and reach the folded state with almost identical rates. The high energy barrier that synchronizes folding is thus conserved. The strong differences in stability between these proteins develop only after this barrier and lead to major changes in the unfolding rates. We conclude that the refolding mechanism of (βα)(8)-barrel proteins is robust. It evolved early and, apparently, has remained conserved upon the diversification of sequences and functions that have taken place within this large protein family.
    No preview · Article · Jul 2012 · Journal of the American Chemical Society

Publication Stats

11k Citations
1,139.30 Total Impact Points


  • 1990-2015
    • University of Bayreuth
      • • Bayreuth Center for Molecular Biosciences (BZMB)
      • • Chair of Biochemistry
      Bayreuth, Bavaria, Germany
  • 2002-2008
    • University Hospital Regensburg
      • Klinik und Poliklinik für Herz-, Thorax- und herznahe Gefäßchirurgie
      Ratisbon, Bavaria, Germany
  • 1984-2005
    • Universität Regensburg
      • • Department of Cardiac, Thoracic and Vascular Surgery near the Heart
      • • Institute of Biophysics and physical Biochemistry
      Ratisbon, Bavaria, Germany
  • 1988-2001
    • Johannes Gutenberg-Universität Mainz
      • Department of Cardiothoracic and Vascular Surgery
      Mainz, Rhineland-Palatinate, Germany
  • 1990-1996
    • Deutsche Gesellschaft für Thorax-, Herz- und Gefäßchirurgie e.V. 
      Mayence, Rheinland-Pfalz, Germany
  • 1987-1989
    • Martin Luther University of Halle-Wittenberg
      Halle-on-the-Saale, Saxony-Anhalt, Germany
  • 1979
    • Stanford University
      • Department of Biochemistry
      Stanford, California, United States