Bringing the Heavy Chain to Light: Creating a Symmetric, Bivalent IgG-Like Bispecific

Bispecific molecules are biologically significant, yet their complex structures pose important manufacturing and pharmacokinetic challenges. Nevertheless, owing to similarities with monoclonal antibodies (mAbs), IgG-like bispecifics conceptually align well with conventional expression and manufacturing platforms and often exhibit potentially favorable drug metabolism and pharmacokinetic (DMPK) properties. However, IgG-like bispecifics do not possess target bivalency and current designs often require tedious engineering and purification to ensure appropriate chain pairing. Here, we present a near-native IgG antibody format, the 2xVH, which can create bivalency for each target or epitope and requires no engineering for cognate chain pairing. In this modality, two different variable heavy (VH) domains with distinct binding specificities are grafted onto the first constant heavy (CH1) and constant light (CL) domains, conferring the molecule with dual specificity. To determine the versatility of this format, we characterized the expression, binding, and stability of several previously identified soluble human VH domains. By grafting these domains onto an IgG scaffold, we generated several prototype 2xVH IgG and Fab molecules that display similar properties to mAbs. These molecules avoided the post-expression purification necessary for engineered bispecifics while maintaining a capacity for simultaneous dual binding. Hence, the 2xVH format represents a bivalent, bispecific design that addresses limitations of manufacturing IgG-like bispecifics while promoting biologically-relevant dual target engagement.

Cloning and expression of maltooligosyltrehalose trehalohydrolase from Sulfolobus solfataricus DSM 1616 in Bacillus subtilis WB800

Maltooligosyltrehalose trehalohydrolase (MTHase) is an industrial enzyme for the production of trehalose. A DNA fragment of 1680 bp encoding for MTHase was cloned from Sulfobolus solfataricus DSM 1616 then fused with promoter acoA-amyE already amplified from pMSE3 vector by PCR to generate an expression cassette acoMTH. Afterward the cassette was inserted into pAC7 vector for expression of the gene in Bacillus subtilis WB800 – a conventional expression system. Gene MTH was inserted into the genome of B. subtilis WB800 by cross-exchange event of pAC7 vector with the host genome for expression of high quality and high quantity of extracellular recombinant protein. By crossing-exchange event at 3’amyE-5’amyE, the expressional cassette was integrated into B. subtilis WB800 genome. The expressional cassette was integrated into B. subtilis WB800 genome replacing 3’amyE-5’amyE, hindering the native amylase activity of the host. Expression of expected protein was confirmed by electrophoresis SDS-PAGE. From our results, it indicates that gene MTH was expressed successfully in B. subtilis WB800. After 0.5% acetoin induction for 48 h, the data showed that the protein with a molecular mass of ~64 kDa on SDS-PAGE was expressed. The level of recombinant protein in WBpAacoMTH was increased and reached 2.5%, 15.2% and 21.95%, respectively comparing with native B. subtilis WB800.

Centromere identity and function put to use: construction and transfer of mammalian artificial chromosomes to animal models

Mammalian artificial chromosomes (MACs) are widely used as gene expression vectors and have various advantages over conventional expression vectors. We review and discuss breakthroughs in MAC construction, initiation of functional centromeres allowing their faithful inheritance, and transfer from cell culture to animal model systems. These advances have contributed to advancements in synthetic biology, biomedical research, and applications in industry and in the clinic.

Selenolysine: A New Tool for Traceless Isopeptide Bond Formation

<p>Despite their biological importance, post-translationally modified proteins are notoriously difficult to produce in a homogeneous fashion using conventional expression systems. Chemical protein synthesis or semi-synthesis offers a solution to this problem; however, traditional strategies often rely on sulfur-based chemistry that is incompatible with the presence of multiple cysteine residues in the target protein. To overcome these limitations, we present the design and synthesis of γ-selenolysine, a selenol-containing form of the commonly modified proteinogenic amino acid, lysine. The utility of γ-selenolysine is demonstrated with the traceless ligation of the small ubiquitin-like modifier protein, SUMO-1, to a peptide segment of human glucokinase. The resulting polypeptide is poised for native chemical ligation and selective deselenization in the presence of unprotected cysteine residues. Selenolysine’s straightforward synthesis and incorporation into synthetic peptides marks it as a universal handle for conjugating any ubiquitin-like modifying protein to its target.</p>

Selenolysine: A New Tool for Traceless Isopeptide Bond Formation

<p>Despite their biological importance, post-translationally modified proteins are notoriously difficult to produce in a homogeneous fashion using conventional expression systems. Chemical protein synthesis or semi-synthesis offers a solution to this problem; however, traditional strategies often rely on sulfur-based chemistry that is incompatible with the presence of multiple cysteine residues in the target protein. To overcome these limitations, we present the design and synthesis of γ-selenolysine, a selenol-containing form of the commonly modified proteinogenic amino acid, lysine. The utility of γ-selenolysine is demonstrated with the traceless ligation of the small ubiquitin-like modifier protein, SUMO-1, to a peptide segment of human glucokinase. The resulting polypeptide is poised for native chemical ligation and selective deselenization in the presence of unprotected cysteine residues. Selenolysine’s straightforward synthesis and incorporation into synthetic peptides marks it as a universal handle for conjugating any ubiquitin-like modifying protein to its target.</p>

Buta Aksara إلى الكلمة Anjing Hutanتأثير ترجمة الأسماء المركبة من معجم اللغة الإندونيسية الكبير على لغة متعلمي اللغة العربية من الكلمة

This study focused on translating the correct compound words into Arabic that become a problem from the Arabic learners as they translate these words into literal translation. The way in which this study was carried out was based on the scientific approach: the descriptive analytical approach for the analysis of the correct translation of the compound words, the method of testing and resolution to learn the effect of translating compound words on Arabic learners and the resolution was composed of nine axes. This study has shown a number of results, the most important of which are: compound words are a set of two or more words that have the same meaning, analysis of the correct translation of the compound words from (KBBI) into three sections, namely: Compound words that were Contrary to the composition of the Arabic vocabulary, of compound words in the type of conventional expression compound words that correspond to the composition of the Arabic vocabulary, and The field study found that the correct translation of the compound words increases the terminology and vocabulary of students to develop the competency level of their language, The correct translation of compound words gives positive effect for development of Arabic learners.

Participating Contingent Premium Options

The first motivation of the creation of derivatives is hedging risk but unfortunately this motivation has changed over the decades since more conventional contracts are used for speculation. The purpose of this study is to use derivatives solely for hedging while respecting principles of profit and risk sharing. According to previous work about the pricing of Waad Bil Mourabaha and using the conventional expression of the contingent premium option, we will propose a model of Participating CPO.