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21 p.-9 fig.-2 tab., Focal adhesion kinase (FAK) is a key component of the membrane proximal signaling layer in focal adhesion complexes, regulating important cellular processes, including cell migration, proliferation, and survival. In the cytosol, FAK adopts an autoinhibited state but is activated upon recruitment into focal adhesions, yet how this occurs or what induces structural changes is unknown. Here, we employ cryo-electron microscopy to reveal how FAK associates with lipid membranes and how membrane interactions unlock FAK autoinhibition to promote activation. Intriguingly, initial binding of FAK to the membrane causes steric clashes that release the kinase domain from autoinhibition, allowing it to undergo a large conformational change and interact itself with the membrane in an orientation that places the active site toward the membrane. In this conformation, the autophosphorylation site is exposed and multiple interfaces align to promote FAK oligomerization on the membrane. We show that interfaces responsible for initial dimerization and membrane attachment are essential for FAK autophosphorylation and resulting cellular activity including cancer cell invasion, while stable FAK oligomerization appears to be needed for optimal cancer cell proliferation in an anchorage-independent manner. Together, our data provide structural details of a key membrane bound state of FAK that is primed for efficient autophosphorylation and activation, hence revealing the critical event in integrin mediated FAK activation and signaling at focal adhesions., I.A. acknowledges support from the José Castillejo Mobility Grant issued by the Spanish Ministry of Education, Culture and Sports. R.D.R. acknowledges funding from the Fellowships for Excellence program sponsored by the Werner-Siemens Foundation and the University of Basel.H.S. acknowledges support by the Swiss National Science Foundation(grants 205320_166164 and 185544 (NCCR TransCure)). M.C.F., A.B., J.C., and C.S. were supported by a Cancer Research UK Programme Grant (C157/ A24837) to M.C.F. F.G. acknowledges funding by the Klaus Tschira Foundation, by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Germany’s Excellence Strategy – 2082/1 –390761711, and the state of Baden-Württemberg through bwHPC and the DFG through grant INST 35/11341 FUGG.S.d.B. thanks the Carl Zeiss Foundation for financial support. D.L. acknowledges support from the Spanish Ministry of Economy, Industry and Competitiveness for the Retos Grant BFU2016-77665-R and the Ministry of Science, Innovation and Universities for the Spanish State Research Agency Retos Grant RTI2018-099318-B-I00,both co-funded by the European Regional Development Fund (FEDER) and from an intramural grant from the Spanish National Research Council(CSIC, Ref: 201820I124). O.L. is funded by SAF2017-82632-P grant from the Spanish Ministry of Science, Innovation and Universities (MCIU/AEI), cofunded by the European Regional Development Fund (ERDF), by the National Institute of Health Carlos III and by projects Y2018/BIO4747 and P2018/NMT4443 from the Autonomous Region of Madrid and co-funded by the European Social Fund and the European Regional Development Fund. C.F.R. is funded by a BES-2015-071348 PhD fellowship by the Spanish Ministry of Science, Innovation and Universities (MCIU/AEI).

Q88Y25_Lacpl is an esterase produced by the lactic acid bacterium Lactobacillus plantarum WCFS1 that shows amino-acid sequence similarity to carboxylesterases from the hormone-sensitive lipase family, in particular the AFEST esterase from the archaeon Archaeoglobus fulgidus and the hyperthermophilic esterase EstEI isolated from a metagenomic library. N-terminally His6-tagged Q88Y25_Lacpl has been overexpressed in Escherichia coli BL21 (DE3) cells, purified and crystallized at 291 K using the hanging-drop vapour-diffusion method. Mass spectrometry was used to determine the purity and homogeneity of the enzyme. Crystals of His6-tagged Q88Y25_Lacpl were prepared in a solution containing 2.8 M sodium acetate trihydrate pH 7.0. X-ray diffraction data were collected to 2.24 A ¿ resolution on beamline ID29 at the ESRF. The apparent crystal point group was 422; however, initial global analysis of the intensity statistics (data processed with high symmetry in space group I422) and subsequent tests on data processed with low symmetry (space group I4) showed that the crystals were almost perfectly merohedrally twinned. Most probably, the true space group is I4, with unit-cell parameters a = 169.05, b = 169.05, c = 183.62 A, JMM and RM thank the Ministerio de Educación y Ciencia for research grants (BFU2010-17929/BMC, AGL2008-01052 and AGL2011-22745; DGCYT), ‘Factoría de Cristalización’ (CSD2006-00015) and FUN-C-FOOD (CSD2007-00063) Consolider-Ingenio 2010 and ALIBIRD S2009/AGR-1469 (CAM) and RM2008-00002 (INIA) for support of their research.

Tannase is an enzyme with important biotechnological applications in the food industry. Previous studies have identified the tannase encoding gene in Lactobacillus plantarum and also have reported the description of the purification of recombinant L. plantarum tannase through a protocol involving several chromatographic steps. Here, we describe the high-yield production of pure recombinant tannase (17 mg/L) by a one-step affinity procedure. The purified recombinant tannase exhibits optimal activity at pH 7 and 40 C. Addition of Ca2þ to the reaction mixture greatly increased tannase activity. The enzymatic activity of tannase was assayed against 18 simple phenolic acid esters. Only esters derived from gallic acid and protocatechuic acid were hydrolyzed. In addition, tannase activity was also assayed against the tannins tannic acid, gallocatechin gallate, and epigallocatechin gallate. Despite L. plantarum tannase representing a novel family of tannases, which shows no significant similarity to tannases from fungal sources, both families of enzymes shared similar substrate specificity range. The physicochemical characteristics exhibited by L. plantarum recombinant tannase make it an adequate alternative to the currently used fungal tannases, This work was supported by grants AGL2005-00470, AGL2008-01052, BFU2007-67404/BMC, Consolider INGENIO 2010 CSD2007-00063 FUN-C-FOOD, and Consolider INGENIO 2007 Factoria de Crystalización (CICYT), RM2008-00002 (INIA), and S-0505/AGR/000153 (CAM). H. Rodríguez and J. A. Curiel were recipients of predoctoral fellowships from the I3P-CSIC Program and FPI-MEC, respectively.