Purification and characterization of beta-glucosidase of Alcaligenes faecalis

A cellobiose-utilizing bacterium isolated from sugar cane bagasse and identified as a strain of Alcaligenes faecalis (ATCC 21400) produced an inducible beta-glucoside-splitting enzyme. The enzyme was purified by a series of streptomycin and ammonium sulfate fractionations and by Sephadex and diethylaminoethyl column chromatography. The final preparation was purified 130-fold, with a recovery of about 10% of the initial enzyme activity. The enzyme had a wide pH range, with optimal activity at pH 6.0 to 7.0. The enzyme was stable in solution at pH 6.5 to 7.8 when kept at 30 C for 2 hr, but it was destroyed by temperatures above 55 C. At 58 and 60 C, the time required to inactivate 90% of the initial activity was 16 and 6.5 min, respectively. An activation energy of 9,500 cal/mole and a K(m) of 1.25 x 10(-4)m were obtained by using p-nitrophenyl beta-glucoside as a substrate. The K(i) value and hydrolysis of cellobiose by the enzyme indicated a high affinity of the enzyme for the cellobiose. The enzyme had its specificity on beta-glucosidic linkage and the rate of hydrolisis of glucosides depended upon the nature of the aglycon moiety. The inactivation studies showed the presence of sulfhydryl groups in the enzyme. The activity of the enzyme was easily destroyed by the Cu(++) and Hg(++) ions. The Michaelis-Menton relationship and the rate of heat inactivation indicated the presence of one type of noninteracting active site in the bacterial beta-glucosidase. Molecular weight of the enzyme was estimated by gel filtration (Sephadex G-200) and sucrose density gradient, and a value of 120,000 to 160,000 was obtained.     

Isolation and Characterization of a Cellulose-utilizing Bacterium

A cellulose-decomposing aerobic and mesophilic bacterium has been isolated from soils of sugar cane fields. The terminal dilution method was adapted to isolate a single clone of cellulolytic organism from closely related contaminants. The cultural and physiological characteristics of the isolate were studied, and the organism was identified as a member of the genus Cellulomonas. The isolate excreted cellulase into the menstruum, and it hydrolyzed various cellulosic materials producing cellobiose as the final breakdown product in the menstruum. When sugar cane bagasse was properly treated with alkali and heat, the organism could decompose up to 90% of the initial substrate within 5 days. Amino acid analysis of the cell crop revealed a high content of lysine, and the essential amino acid pattern compared favorably with that of Food and Agricultural Organization reference protein.     

小麦再生植株的变异研究 Ⅱ.再生植株当代(R_1代)的细胞学和形态学变异

再生植株具有高频率的染色体异常,其中有20.61％表现为染色体数量变异,最常见的为2n—1类型,其次为2n—2类型,也有2n 1、2n—3个体以及染色体数嵌合株。再生植株减数分裂各期均有染色体异常行为,可以见到的有落后染色体、染色体桥、断片、二分体延迟、微核,还有粗线期十字型配对等结构变异,以及五分体、六分体和畸型四分体等异常现象。微核率随培养时间延长而增加,可用作染色体伤害的一个指标。再生植株R_1代存在着许多形态学变异。性状变异与染色体数目变异没有明显关系。     

Inositol 1,4,5-trisphosphate-induced calcium release in the organelle layers of the stratified, intact egg of Xenopus laevis

Using double-barreled, Ca2(+)-sensitive microelectrodes, we have examined the characteristics of the Ca2+ release by inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) in the various layers of Xenopus laevis eggs in which the organelles had been stratified by centrifugation. Centrifugation of living eggs stratifies the organelles yet retains them in the normal cytoplasmic milieu. The local increase in intracellular free Ca2+ in each layer was directly measured under physiological conditions using theta-tubing, double-barreled, Ca2(+)-sensitive microelectrodes in which one barrel was filled with the Ca2+ sensor and the other was filled with Ins(1,4,5)P3 for microinjection. The two tips of these electrodes were very close to each other (3 microns apart) enabling us to measure the kinetics of both the highly localized intracellular Ca2+ release and its subsequent removal in response to Ins(1,4,5)P3 injection. Upon Ins(1,4,5)P3 injection, the ER-enriched layer exhibited the largest release of Ca2+ in a dosage-dependent manner, whereas the other layers, mitochondria, lipid, and yolk, released 10-fold less Ca2+ in a dosage-independent manner. The removal of released Ca2+ took place within approximately 1 min. The sensitivity to Ins(1,4,5)P3 and the time course of intracellular Ca2+ release in the unstratified (unactivated) egg is nearly identical to that observed in the ER layer of the stratified egg. Our data suggest that the ER is the major organelle of the Ins(1,4,5)P3-sensitive Ca2+ store in the egg of Xenopus laevis.     

Virological survey of rhesus monkeys in China

A virological survey of rhesus monkeys captured in China for 13 viruses and/or antibodies was performed. Antigens used were SFV, SF40, HSV-1, Sa11, measles, vaccinia, epidemic or simian hemorrhagic fever, Langat, Kunming, poliomyelitis, HIV, SV41 and rubella. Monkeys were from Sichuan, Hunan, Guizhou, Yunnan and Guangxi provinces. Antibody was detected to all the listed viruses except HIV, SV41 and rubella. Both SFV and SV40 were recovered from monkeys, but H. simiae, LCM and coxsackieviruses were not.     

大肠杆菌青霉素G酰化酶基因及其邻近区域的核苷酸全序列

Some of microorganisms have been known to possess penicillin G acylase activity. The E. coli derived penicillin G acylase (PGA) can catalyze the conversion of penicillin G into phenylacetic acid and 6-amino-penicillanic acid, the latter is used as the starting compound for the industrial formation of semi-synthetic penicillins. Apart from its industrial importance, the enzyme PGA displays a number of interesting properties. Catalytically active enzyme is localized in the periplasmic space of E. coli cells and composed of two dissimilar subunits. The two subunits are apparently produced from a precursor protein, via a processing pathway hitherto unique in its features for a prokaryotic enzyme. The studies on processing of the precursor and on the relationship between structure and function of the mature enzyme are important theoretically. Previously we cloned a 3.5 kb DNA fragment from a strain (E. coli AS 1.76), which displays PGA activity. In this paper, we report a nucleotide sequence of the 3.5 kb DNA fragment containing PGA gene. After insertion of the DNA fragment into EcoR I and Hind III sites in pWR 13, pPGA 20 had been obtained. We subcloned the Hind III and Bg1 II treated fragment of 1.6 kb in length from pPGA 20 into Hind III and BamH I sites of pWR 13 to get a pPGA 1.6, and Bg1 II and EcoR I treated fragment of 1.9 kb in length into BamH I and EcoR I sites of pWR 13 to get a pPGA 1.9. The linearized pPGA 1.9 which were digested with appropriate restriction enzymes were progressively shortened from both ends respectively by digestion with Bal 31 nuclease, followed by cleavage of shortened target DNA off vector DNA molecules with appropriate restriction enzymes. The series of the DNA fragments shortened from EcoR I end were then cloned into plasmid pWR 13 which had previously digested with Hind III and Sma I enzymes (Fig. 1). The DNA fragment cloned in pWR 13 were directly sequenced on the resulted plasmids by using primer I and primer II. Thus we have obtained the complete nucleotide sequence of the 3.5 kb DNA fragment. The 3.5 kb fragment contains an intact PGA gene which is 2.6 kb.(ABSTRACT TRUNCATED AT 400 WORDS)     

A neural network algorithm for the multiple traveling salesmen problem

We developed an efficient neural network algorithm for solving the Multiple Traveling Salesmen Problem (MTSP). A new transformation of the N-city M-salesmen MTSP to the standard Traveling Salesmen Problem (TSP) is introduced. The transformed problem is represented by an expanded version of Hopfield-Tank's neuromorphic city-position map with (N + M-1)-cities and a single fictitious salesmen. The dynamic model associated with the problem is based on the Basic Differential Multiplier Method (BDMM) [26] which evaluates Lagrange multipliers simultaneously with the problem's state variables. The algorithm was successfully tested on many problems with up to 30 cities and five salesmen. In all test cases, the algorithm always converged to valid solutions. The great advantage of this kind of algorithm is that it can provide solutions to complex decision making problems directly by solving a system of ordinary differential equations. No learning steps, logical if statements or adjusting of parameters are required during the computation. The algorithm can therefore be implemented in hardware to solve complex constraint satisfaction problems such as the MTSP at the speed of analog silicon VLSI devices or possibly future optical neural computers.