• 대한방사성의약품학회지
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• 제7권1호
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• pp.33-40
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• 2021

Recently, antibody-drug conjugates (ADCs) are used to deliver efficient cytotoxic payloads selectively in cancer cells. In the designing of an ADC, the antibody is connected to a toxic payload via a covalent linker, which helps to solubilizes the typical hydrophobic payload as well as stabilizes the linkage over circulation. The development of the linkers for the antibody drug conjugate is still in demand. Initially, the acid, disulfide, and cathepsin-sensitive ADCs attracted considerable attention for the delivery of a potent cytotoxic payload but suffer from instability in human and mouse plasma with a short half-life. In addition, It also suffer from a solubility issue that induces aggregation, which is the major problem in their development. ADCs associated with sulfatase and phosphatase cleavable linker are highly soluble due to the anionic nature of sulfate and phosphate groups. The ADCs also showed high stability in human and mouse plasma. Therefore, to overcome these limitations, sulfatase and phosphatase cleavable linkers were developed. This review focuses on the recently reported advantages of sulfatase and phosphatase cleavable linkers for ADCs.

• 산업기술연구
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• 제36권
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• pp.33-37
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• 2016

Cellulose binding domains (CBDs) of cellulases are thought to assist in the hydrolysis of insoluble crystalline cellulose. To gain sufficient amount of CBDs, the self-cleavable intein tag was used for expression and purification of Trichoderma reesei cellobiohydrolase I CBD in E. coli. Synthetic CBD genes, CBD or linker-CBD were cloned into expression vector pTYB11. Recombinant CBDs were successfully purified by intein mediated purification with an affinity chitin-binding domain. The final yields of recombinant CBD and linker-CBD were 3.2 mg/L and 1.4 mg/L, respectively. The functional bindings of recombinant CBDs were confirmed by Avicel binding experiments. The simple and easy purification method using self-cleavable intein tag can be further used in pretreatment of crystalline cellulose or characterization of engineered CBDs.

• Fisheries and Aquatic Sciences
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• 제4권2호
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• pp.84-87
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• 2001

Multimeric angiotensin-converting-enzyme-inhibitor (ACE}) containing a trypsin cleavable linker peptide between ACEI was constructed. We made synthetic DNA coding for the ACEI peptide with asymmetric and complementary cohesive ends of linker nucleotides. A tandemly repeated DNA cassette for the expression of concatameric short peptide multimers was constructed by ligating the basic units. The resultant multimeric peptide expressed as soluble and trypsin treated peptide was shown at the same retention time with chemically synthetic ACEI by HPLC. The present results showed that the technique developed for the production of the ACEI multimers with trypsin cleavable linker peptides can be generally applicable to the production of short peptide.

• Journal of Microbiology and Biotechnology
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• 제33권12호
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• pp.1681-1691
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• 2023

Flavin mononucleotide-binding proteins or domains emit cyan-green fluorescence under aerobic and anaerobic conditions, but relatively low fluorescence and less thermostability limit their application as reporters. In this work, we incorporated the codon-optimized fluorescent protein from Chlamydomonas reinhardtii with two different linkers independently into the redox-responsive split intein construct, overexpressed the precursors in hyperoxic Escherichia coli SHuffle T7 strain, and cyclized the target proteins in vitro in the presence of the reducing agent. Compared with the purified linear protein, the cyclic protein with the short linker displayed enhanced fluorescence. In contrast, cyclized protein with incorporation of the long linker including the myc-tag and human rhinovirus 3C protease cleavable sequence emitted slightly increased fluorescence compared with the protein linearized with the protease cleavage. The cyclic protein with the short linker also exhibited increased thermal stability and exopeptidase resistance. Moreover, induction of the target proteins in an oxygen-deficient culture rendered fluorescent E. coli BL21 (DE3) cells brighter than those overexpressing the linear construct. Thus, the cyclic reporter can hopefully be used in certain thermophilic anaerobes.

• Biomolecules & Therapeutics
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• 제5권1호
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• pp.53-58
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• 1997

$[Lys^7$]dermorphin, abbreviated K7DA, which has structural features similar to a metabolically stable $\mu$-opioid peptide agonist $[D-Arg^2, Lys^4$]dermorphin analogue (DALDA), but is intrinsically more potent with respect to binding to the $\mu$-opioid peptide receptor. The present studies report on attempts to enhance brain uptake of systemically administered K7DA by conjugation to a complex of streptavidin (SA) and the OX26 murine monoclonal antibody to the rat transferrin receptor, which undergoes receptor-mediated transcytosis through the blood-brain barrier (BBB). SA-OX26 conjugate mediates BBB transport of biotinylated therapeutics. The K7DA is monobiotinylated at the $\varepsilon$-amino group of the $[Lys^7$] residue with cleavable linker using NHS-SS-biotin. The brain uptake of $^{125}I$ labeled biotinylated K7DA ($^{125}I$-bio-SSa-K7DA) was very small and rapidly metabolized after intravenous injection. The brain uptake, expressed as percent of injected dose delivered per gram of brain, of the $^{125}I$-bio-55-K7DA bound to the SA-OX26 conjugate $^{125}I$-bio-SS-K7DA/SA-OX26) was 0.14$\pm$0.01, a level that is 2-fold greater than the brain uptake of morphine. The cleavability of the disulfide linker in vivo in rat plasma and brain was assessed with gel filtration HPLC and intravenous injection of labeled opioid chimeric peptides. The disulfide linker is stable in plasma in vivo but is cleaved in rat brain in vivo. In conclusion, these studies show that delivery of these potential opioid peptides to the brain may be improved by coupling them to vector-mediated BBB drug delivery system.

• 한국식품위생안전성학회지
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• 제18권3호
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• pp.101-106
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• 2003

냉장식품의 주요한 변패원일균으로서 저온세균인 Pseudomonas aeruhinosa를 최소 전처리한 후 신속히 검출할 수 있는 비표지 면역센서 시스템을 개발하였다. 수정결정전극상으로의 생물요소인 항체의 고정화는 이형이기능성 가교화제인 sulfosuccinimidyl 6-[3-(2-pyridyldithio)propionamido] hexanoate를 사용하여 항체를 티올화시킨 후 티올화된 항체를 화학흡착하여 행하였고, P. aeruginosa flagella에 대한 단클론항체를 사용하였을 때 다클론항체를 사용한 경우보다 센서감응이 우수한 것으로 나타났다. 항체가 고정화된 센서 chip과 flow형 quart crystal microbalance 계측 시스템을 이용하여 균 첨가 및 증균을 행한 10종의 모델시료에 대한 계측을 행하였다. 이 때, 시료자체에 의한 진동수변화가 52~89 Hz 범위인 반면 균 첨가 시에 나타난 진동수변화는 108~200 Hz이었고 증균시료에 의한 진동수변화는 162~222 Hz 범위로 나타났다. 시스템 안정화, 시료주입 및 정상상태이 센서반응 획득, 시스템 세척으로 이루어지는 한 주기의 센서계측에 소요된 시간은 모든 시료에 있어 30분 이내였다.