Anchoring of Surface Proteins to the Cell Wall of Staphylococcus aureus. III. Lipid II is an in vivo peptidoglycan substrate for sortase-catalyzed surface protein anchoring

Surface proteins of Staphylococcus aureus are anchored to the cell wall peptidoglycan by a mechanism requiring a C-terminal sorting signal with an LPXTG motif. Surface proteins are first synthesized in the bacterial cytoplasm and then transported across the cytoplasmic membrane. Cleavage of the N-terminal signal peptide of the cytoplasmic surface protein P1 precursor generates the extracellular P2 species, which is the substrate for the cell wall anchoring reaction. Sortase, a membrane-anchored transpeptidase, cleaves P2 between the threonine (T) and the glycine (G) of the LPXTG motif and catalyzes the formation of an amide bond between the carboxyl group of threonine and the amino group of cell wall cross-bridges. We have used metabolic labeling of staphylococcal cultures with [32P]phosphoric acid to reveal a P3 intermediate. The 32P-label of immunoprecipitated surface protein is removed by treatment with lysostaphin, a glycyl-glycine endopeptidase that separates the cell wall anchor structure. Furthermore, the appearance of P3 is prevented in the absence of sortase or by the inhibition of cell wall synthesis. 32P-Labeled cell wall anchor species bind to nisin, an antibiotic that is known to form a complex with lipid II. Thus, it appears that the P3 intermediate represents surface protein linked to the lipid II peptidoglycan precursor. The data support a model whereby lipid II-linked polypeptides are incorporated into the growing peptidoglycan via the transpeptidation and transglycosylation reactions of cell wall synthesis, generating mature cell wall-linked surface protein

Anchoring of Surface Proteins to the Cell Wall of Staphylococcus aureus: sortase catalyzed in vitro transpeptidation reaction using LPXTG peptide and NH2-Gly3 substrates

Staphylococcus aureus sortase anchors surface proteins to the cell wall envelope by cleaving polypeptides at the LPXTG motif. Surface proteins are linked to the peptidoglycan by an amide bond between the C-terminal carboxyl and the amino group of the pentaglycine cross-bridge. We find that purified recombinant sortase hydrolyzed peptides bearing an LPXTG motif at the peptide bond between threonine and glycine. In the presence of NH2-Gly3, sortase catalyzed exclusively a transpeptidation reaction, linking the carboxyl group of threonine to the amino group of NH2-Gly3. In the presence of amino group donors the rate of sortase mediated cleavage at the LPXTG motif was increased. Hydrolysis and transpeptidation required the sulfhydryl of cysteine 184, suggesting that sortase catalyzed the transpeptidation reaction of surface protein anchoring via the formation of a thioester acyl-enzyme intermediate

Transfer Learning from Audio Deep Learning Models for Micro-Doppler Activity Recognition

This paper presents a mechanism to transform radio micro-Doppler signatures into a pseudo-audio representation, which results in significant improvements in transfer learning from a deep learning model trained on audio. We also demonstrate that transfer learning from a deep learning model trained on audio is more effective than transfer learning from a model trained on images, which suggests machine learning methods used to analyse audio can be leveraged for micro-Doppler. Finally, we utilise an occlusion method to gain an insight into how the deep learning model interprets the micro-Doppler signatures and the subsequent pseudo-audio representations

Perencanaan Jembatan Pelengkung Type Throught Arch Desa Korek, Kec. Ambawang, Kab. Kubu Raya (Provinsi Kalimantan Barat)

Jembatan merupakan suatu struktur bangunan yang berfungsi untuk menyatukan jalan yang terputus oleh rintangan, misalnya sungai, rawa, dll. Dalam penyusunan Tugas Akhir ini direncanakan jembatan pelengkung type Throught Arch di Desa Korek, Kec. Ambawang, Kab. Kubu Raya (Ka-Bar). Tahap awal perencanaan adalah perhitungan struktur atas yang meliputi pipa sandaran, tiang sandaran, dinding penahan, trotoar, pelat lantai kendaraan, gelegar memanjang, gelegar melintang, hubungan gelegar memanjang terhadap gelegar melintang dan sekaligus perhitungan penghubung geser (shear connector). Memasuki tahap konstruksi utama, dilakukan perhitungan beban-beban yang bekerja kemudian dianalisa dengan menggunakan program komputer. Setelah didapat gaya-gaya dalam yang bekerja dilakukan perhitungan kontrol tegangan dan perhitungan sambungan. Bersamaan dilakukan perhitungan konstruksi pemikul utama juga dilakukan perhitungan konstuksi sekunder yang meliputi ikatan angin bawah, ikatan angin busur atas dan ikatan angin busur bawah. Dari hasil perencanaan didapat profil yang dipakai dalam perencanaan jembatan pelengkung type throught arch di Desa korek, Kec. Ambawang, Kab. Kubu Raya (KALBAR), yaitu: Gelegar Memanjang IWF 400 x 300 x 16 x 10, Gelegar Melintang IWF 700 nx 300 x 28 x 15, Ikatan Angin Bawah, Ikatan Angin Busur Atas dan Ikatan Angin Busur Bawah IWF 150 x 150 x 10 x 7, Busur Atas 30\u27\u27 WF 30 x 15, Busur Bawah 30\u27\u27 WF 30 x 15, Gelegar Induk Bagian Tepi 30\u27\u27 WF 30 x 15, Batang Penggantung IWF 200 x 200 x 12 x 8, Batang Tegak IWF 200 x 200 x 12 x 8, Batang Diagonal IWF 200 x 200 x 12 x 8, Batang Tegak Bagian Tepi 30\u27\u27 WF 30 x 15 dan Batang Diagonal 30\u27\u27 WF 30 x 15

White Gaussian Noise Based Capacity Estimate and Characterization of Fiber-Optic Links

We use white Gaussian noise as a test signal for single-mode and multimode transmission links and estimate the link capacity based on a calculation of mutual information. We also extract the complex amplitude channel estimations and mode-dependent loss with high accuracy.Comment: submitted to The Optical Networking and Communication Conference (OFC) 201

Contribution analysis of a Bolivian innovation grant fund: mixing methods to verify relevance, efficiency and effectiveness

We used contribution analysis to verify the key assumption in the intervention logic of an innovation fund in Bolivia directed to economic farmer organisations to develop value-added activities. We focused the research on three sub-components of the intervention logic: relevance of the farmer groups for local economic development, effectiveness of the fund in strengthening these group, and efficiency of the grant allocation mechanism. We used a case-based comparative analysis to assess effectiveness: improved market access for members, strengthened organisational capacities and the capacity to pay organisational costs. We showed that the grants to already well-endowed organisations were particularly unsuccessful