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With the development of recombinant DNA technology, a growing number of peptides and proteins are being applied to clinical treatment of diseases. However, a huge challenge needed to be addressed for many therapeutic peptides or proteins is that their half-lives in circulation is short due to rapid renal clearance and enzymatic degradation. To overcome the challenge, half-life extension strategies have been extensively developed. Albumin-binding strategy, whether covalent or non-covalent, represent a widely applied and highly successful half-life extension strategy, and many products have been marketed or tested in clinical trials. This review focuses on the utilization of albumin for half-life prolongation and emphasizes on the albumin-binding drugs already on the market or in the clinical stage. We summarize the related techniques including genetic fusion, chemical conjugation, non-covalent binding fatty acid and protein moieties.

Immobilization of enzymes is important and widely applied in biocatalysis. Streptomyces platensis gene gox, encoding an extracellular L-glutamate oxidase (Gox), was fused to cellulose binding domain (CBDcex) from Cellulomonas fimi and the recombinant protein Gox-CBD was expressed in Escherichia coli. The fusion protein (Gox-CBD) was immobilized onto microcrystalline cellulose. The preparation conditions, binding capacity, properties and stability of the immobilized enzyme were studied. Under the condition of 4 ℃, for 1 hour, the fusion protein Gox-CBD was able to bind microcrystalline cellulose at a ratio of 9.0 mg of protein per gram of microcrystalline cellulose. Enzymatic properties of free and immobilized L-glutamic oxidase (Gox-CBD) were compared. The specific activity of the immobilized enzyme decreased, but its thermal stability increased a lot compared with that of the free Gox-CBD. After incubation at 60 ℃ for 30 min, 70% of the total activity remained whereas the free recombinant Gox completely lost its activity. The immobilized protein was tightly bound to microcrystalline cellulose at pH below 10 or more than 5 mmol/L NaCl. The fusion protein of Gox-CBD can be specifically immobilized on the microcrystalline cellulose on a single step. Therefore, our findings can provide a novel strategy for protein purification and enzyme immobilization.酶的固定化作为一种重要的技术，已在生物催化领域得到了广泛的应用。现将来源于普拉特链霉菌3304 (Streptomyces platensis NTU3304) 产生的胞外L-谷氨酸氧化酶 (L-glutamate oxidase，Gox) 基因gox 融合到来源于粪碱纤维单胞菌Cellulomonas fimi 的纤维素结合域 (CBDcex) 的基因上， 构建表达载体pETM10-Gox-CBD，并在大肠杆菌中表达。通过蛋白纯化获得融合蛋白，并命名为Gox-CBD。利用CBD 对微晶纤维素特异性吸附的特性将其固定在微晶纤维素上，并对固定化酶的制备条件、结合量、酶学性质及其微晶纤维素结合稳定性等进行了研究。在4 ℃条件下结合约1 h，融合蛋白Gox-CBD 结合在纤维素上的结合量即可达到9.0 mg/g。通过对重组型、融合表达游离的以及固定化在微晶纤维素上的谷氨酸氧化酶的酶学性质进行比较发现，固定化酶的比酶活有所降低；但固定化酶的热稳定性相对于游离酶有了很大的提高，在60 ℃孵育30 min 后还保留有约70%的活性，而游离的重组Gox 在相同条件下几乎完全失去活性。当固定化结合蛋白在pH10 或者盐浓度5 mmol/L 的NaCl 条件下可以牢固结合。并且可以通过一步纯化方法固定化融合蛋白Gox-CBD 于微晶纤维素上。因此，L-谷氨酸氧化酶与纤维素结合域融合表达的研究为蛋白的纯化及酶的固定化提供了一种新策略。.

The herbicide chlorimuron-ethyl has been applied widely for weed control in farmland, especially in soybean fields in China over the past decade, but the chronic effects of this herbicide on soil microorganisms, particularly Pseudomonas spp., is not well understood. Taking a continuously cropped soybean field in the town of Fuyuan—a soybean production base of Heilongjiang Province in Northeast China—as a case study, soil samples were collected from plots having received 0-, 5-, and 10-year applications of chlorimuron-ethyl (30 g active component of chlorimuron-ethyl/ha/year) to study the abundance and diversity of Pseudomonas spp. Meanwhile, an in vitro assay was used to examine the antifungal activities of isolated Pseudomonas spp. against soil-borne pathogens (Fusarium graminearum, Fusarium oxysporum, and Rhizoctonia solani) causing soybean root rot disease. The production of siderophore, hydrogen cyanide (HCN), and lytic enzymes (cellulase, pectinase, and chitinase) by Pseudomonas spp. was also investigated. With 5- and 10- year chlorimuron-ethyl application, the numbers of soil Pseudomonas spp. decreased from 121 × 102 CFU/g dry soil in the control to 40 × 102 CFU/g dry soil and 13 × 102 CFU/g dry soil, and the Shannon index values decreased from 6.23 to 3.71 and 1.73, respectively. The numbers of antifungal Pseudomonas spp. also decreased, and the proportions of Pseudomonas spp. with antifungal activities against the different test pathogens altered. All the antifungal Pseudomonas spp. could produce siderophore and HCN but not lytic enzymes. The results suggest that long-term application of chlorimuron-ethyl in continuously cropped soybean field had negative effects on the abundance and diversity of soil Pseudomonas spp., including species with different antifungal activities against pathogens. Siderophore and HCN rather than lytic enzymes formed the antifungal metabolites of Pseudomonas spp., and the number of antifungal Pseudomonas that can produce siderophore and HCN decreased markedly under application of chlorimuron-ethyl, especially after 10-year application.

Chlorimuron-ethyl (CE) has been widely used in modern agriculture, but little is known regarding the influence of CE on ammonia-oxidizing bacteria (AOB) and archaea (AOA) populations in soils. In this study, microcosm incubation of aquic brown soil was conducted for 60 d. Associated changes in the population sizes of AOB and AOA in response to CE application with or without urea fertilization were examined via quantitative real-time PCR (qPCR) assays of the ammonia monooxygenase gene (amoA). The half-life of CE ranged from 11.80 d to 14.54 d in the tested soil. Compared to the untreated control, the application of CE alone had no strong effects on soil pH, and urea fertilization temporarily increased soil pH in the first 7 days. The abundance of the AOA amoA gene was greater than the abundance of the AOB amoA gene in all treatments, but both were significantly suppressed by CE application in a dose-dependent manner. Urea fertilization generally increased AOB and AOA amoA gene abundances, except that the AOA amoA gene level was slightly reduced at the early stage of the incubation period. AOB and AOA preferred different N levels for growth, with AOB only growing significantly at high NH4(+) levels and AOA growing substantially at low NH₄(+) levels. The stimulation effects of urea fertilization on AOA and AOB amoA gene abundances were strongly suppressed by the CE application. This study indicated that the CE application substantially suppressed soil nitrification via inhibiting the AOB and AOA population regardless of urea fertilization, which resulted in significant changes in the soil NH₄(+)-N and NO₃(-)-N levels. Furthermore, AOB and AOA inhabiting separate ecological niches with different NH₄(+) levels played various roles in N cycling.