Line geometries for sequence comparisons

Well-known dynamic programming algorithms exist for comparing two finite sequences inO(N ²) time and storage, whereN is the common sequence length. Extensions to the comparison ofM finite sequences requireO((2N) ^M) time and storage, making such algorithms difficult even forM=3. A simple generalization of the sequences makes it possible to obtain some results about the geometry of sequence alignments. These ideas suggest heuristic approaches to problems of comparing several sequences. IfM sequences are known to be related by a binary tree, they can be aligned inO(MN ²) time andO(N ²+NM) storage. This work was supported by a grant from the System Development Foundation.     

Optimal sequence alignment allowing for long gaps

A new algorithm for optimal sequence alignment allowing for long insertions and deletions is developed. The algorithm requires O((L+C)MN) computational steps, O(LN) primary memory and O(MN) secondary memory storage, whereM andN(M≥N) are sequence lengths,L (typicallyL≤3) is the number of segment specifying the gap weighting function, andC is a constant. We have also modified our earlier traceback algorithm so that it finds all and only the optimal alignments in a compact form of a directed graph. The current versions accept a set of aligned sequences as input, which facilitates multiple sequence alignment by some iterative procedures. Dedicated to Professor Akiyoshi Wada on the occasion of his 60th birthday.     

N2O Reduction by Azotobacter vinelandii with Emphasis on Kinetic Nitrogen Isotope Effects

A nitrogen-fixing bacterium Azotobacter vinelandii was successfullygrown in a specially designed system with constant partial pressuresof N₂O (0.2 atm) and O₂ (0.2 atm) in a nitrogen-free liquidmedium. Reduction of N₂O proceeded with the evolution of N₂in the gas phage. Large nitrogen isotope fractionation was found for both processes,reduction of N₂O to N₂ and N₂O-fixation. The kinetic isotopefractionation factors of these reactions were at most 1.039and 1.034, respectively. Furthermore, an unexpected inverseisotope effect (organic-N, the end-product, is more enrichedin ¹⁵N than N₂, the intermediate) strongly suggested that N₂Owas directly assimilated within the bacterial cells. Simultaneousassimilation of N₂O and N₂ was also confirmed by using a ¹⁵Ntracer technique. Three independent pathways were demonstrated for the nitrogenfixing system investigated in this study: (1) a direct reductionof N₂O to ammonium (apparently 8-electron reduction), (2) reductionof N₂ to ammonium (6-electron reduction) and (3) N₂O reductionto N₂ (2-electron reduction). ³ Present address: Department of Environmental Sciences, Facultyof Integrated Arts and Sciences, Hiroshima University, Hiroshima730, Japan ⁴ Present address: Department of Earth Sciences, Faculty ofScience, Toyama University, Gofuku, Toyama 930, Japan (Received June 18, 1986; Accepted December 16, 1986)     

Microheterogeneity of N-glycosylation on a stylar self-incompatibility glycoprotein of Nicotiana alata

Gametophytic self-incompatibility, a mechanism that preventsinbreeding in some families of flowering plants, is mediatedby the products of a single genetic locus, the S-locus. Theproducts of the S-gene in the female sexual tissues of Nicotianaalata are an allelic series of glycoproteins with RNase activity.In this study, we report on the microheterogeneity of N-linkedglycosylation at the four potential N-glycosylation sites ofthe S²-glycoprotein. The S-glycoproteins from N.alata containfrom one to five potential N-glycosylation sites based on theconsensus sequence Asn-Xaa-Ser/Thr. The S²-glycoprotein containsfour potential N-glycosylation sites at Asn²⁷, Asn³⁷, Asn¹³⁸and Asn¹⁵⁰, designated sites I, n, IV and V, respectively. SiteIII is absent from the S₂-glycoprotein. Analysis of glycopeptidesgenerated from the S₂-glycoprotein by trypsin and chymotrypsindigestions revealed the types of glycans and the degree of microheterogeneitypresent at each site. Sites I (Asn²⁷) and IV (Asn¹³⁸) displaymicroheterogeneity, site II (Asn³⁷) contains only a single typeof N-glycan, and site V (Asn¹⁵⁰) is not glycosylated. The microheterogeneityobserved at site I on the S₂-glycoprotein is the same as thatobserved at the only site, site I, on the Srglycoprotein (Woodwardet al., Glycobiology, 2, 241-250, 1992). Since the N-glycosylationconsensus sequence at site I is conserved in all S-glycoproteinsfrom other species of self-incompatible solanaceous plants,glycosylation at this site may be important to their function.No other post-translational modifications (e.g. O-glycosylation,phosphorylation) were detected on the S₂-glycoprotein. fertilization microheterogeneity N-glycans plants RNase     

黔中地区不同植被类型土壤氧化亚氮的释放特征及影响因素

利用密闭箱-气相色谱法于2006—2007年对黔中地区退耕荒草地、灌丛、马尾松林和阔叶林土壤氧化亚氮的释放通量进行原位观测,初步研究了我国南方喀斯特地区不同植被类型土壤N₂O释放量的季节变化.结果表明:除个别月份土壤表现为大气N2O的吸收汇,各观测点均为N₂O的源,植被条件和土壤类型对土壤N₂O的释放具有明显影响.退耕荒草地、灌丛N₂O释放量具有明显的季节变化规律, 春夏季节高于秋冬季节,通量范围分别在-20.7～103.0 μg N·m^-2·h^-1和 -33.0～67.3 μg N·m^-2·h^-1.马尾松林、阔叶林春季土壤N₂O释放量最高,其他季节变化规律不明显,通量范围分别在-5.3～35.0 μg N·m^-2·h^-1和-14.4～152.8 μg N·m^-2·h^-1.相关性分析表明,土壤水分与N₂O释放通量显著负相关,是影响土壤N₂O释放通量季节变化的主要驱动因素.温度通过影响土壤水分而间接影响土壤N₂O的释放通量.     

Yield-density Relationships: the Influence of Resource Availability on Growth and Self-thinning in Populations of Vulpia fasciculata

Monocultures of Vulpia fasciculata were grown over a wide rangeof densities to investigate the influence of crowding and nutrientsupply on growth and self-thinning. For a given time and densityseries the relationship between mean yield per plant (w) andthe density of survivors (N) could be described by the equation w= w_m (1+aN)^–b. where w_m is the yield of an isolated plant, a is the area requiredto achieve a yield of w_m and b describes the effectiveness withwhich resources are taken up from the area. All three parametersincreased with time. Adding nutrients changed not only the rate at which the effectsof crowding occurred but also the intensity of crowding since w_m = C(a^b)^D. where C and D are constants. The addition of nutrients resultedin an increase in the value of C. Such an increase means thata larger weight can be supported by a given area because theresources within that area are greater. During the early phases of growth, populations of V. fasciculataconformed to the –3/2 power law, w = cN^–3/2, butonly at very high densities with a plentiful supply of nutrients.However, once the maximum standing crop had been reached thetrajectory of the thinning line switched to a slope of justless than –1 when weight was ploted against density onlogarithmic scales. The intercept of the –3/2 thinningline was considerably higher (log c = 5.74) than those for mosttrees and forbs but was similar to those of a number of othergrasses. Vulpia fasciculata, dune fescue, yield-density models, self-thinning, density-dependence, nutrient supply     

Proton Translocation Associated with Denitrification in a Photodenitrifier, Rhodopseudomonas sphaeroides forma sp. denitrificans

H⁺ translocation driven by NO₃^–, NO₂^– and N₂O reductionswith endogenous substrates in cells of Rhodopseudomonas sphaeroidesforma sp. denitrificans was investigated by the oxidant pulsemethod. Upon injection of nitrogenous oxides to anaerobic cellsin darkness, an alkaline transient in the external medium wasobserved, followed by acidification. The alkaline transientwas enhanced by carbonyl cyanide m-chlorophenylhydrazone. When a viologen dye was used as an electron donor in the presenceof 1 mM Af-ethylmaleimide and 0.1 mM 2-n-heptyl-4-hydroxyquinoline-N-oxideto preclude respiration-linked H⁺ extrusion, addition of KNO₃,KNO₂ and N₂O caused only a rapid alkalinization. The H⁺ consumptionstoichiometries, H⁺/2e^– ratios for NO₃^– reductionto NO₂^–, NO₂^– reduction to 1/2 N₂O and N₂O reductionto N₂ were –1.90, –3.18 and –2.04, respectively.These values agreed well with the fact that all reductions ofnitrogenous oxides in denitrification occur on the periplasmicside of the cytoplasmic membrane. When corrected for H⁺ consumption in the periplasm, the H⁺ extrusionstoichiometries, H⁺/2e^– ratios with endogenous substratesin the presence of K⁺/valinomycin for NO₃^– reduction toNO₂^–, NO₂^– reduction to 1/2 N₂O and N₂O reductionto N₂ were 4.05, 4.95 and 6.01, respectively. (Received August 4, 1982; Accepted January 13, 1983)     

Effect of Benzyladenine on Grain Growth in aestivum wheat

Effect of benzyladenine (BA) on inter and intraspikelet grainsize spectrum in aestivum wheat (cv. Kalyan Sona) has been comparedwith ¹⁴C-uptake of externally added sucrose by the grains andits incorporation in non-starch endosperm lipid fraction. Itis suggested that BA application enhanced the synthesis of membranestructural components required for storage in endosperm. (Received May 8, 1986; Accepted April 2, 1987)     

Reduced space sequence alignment

Motivation: Sequence alignment is the problem of finding theoptimal character-by-character correspondence between two sequences.It can be readily solved in O(n²) time and O(n²) space on aserial machine, or in O(n) time with O(n) space per O(n) processingelements on a parallel machine. Hirschberg's divide-and-conquerapproach for finding the single best path reduces space useby a factor of n while inducing only a small constant slowdownto the serial version. Results: This paper presents a family of methods for computingsequence alignments with reduced memory that are well suitedto serial or parallel implementation. Unlike the divide-and-conquerapproach, they can be used in the forward-backward (Baum-Welch)training of linear hidden Markov models, and they avoid data-dependentrepartitioning, making them easier to parallelize. The algorithmsfeature, for an arbitrary integer L, a factor proportional toL slowdown in exchange for reducing space requirement from O(n²)to O(n). A single best path member of this algorithm familymatches the quadratic time and linear space of the divide-and-conqueralgorithm. Experimentally, the O(n1.5)-space member of the familyis 15–40% faster than the O(n)-space divide-and-conqueralgorithm. Availability: The methods will soon be incorporated in the SAMhidden Markov modeling package http: //www.cse.ucs-c.edu/research/compbio/sam.html. Contact: wzrph{at}cse.ucsc.edu     

Mutation of Asn128 to Asp of Phaseolus vulgaris leucoagglutinin (PHA-L) eliminates carbohydrate-binding and biological activity

Phytohaemagglutinin (PHA) is the major lectin present in theseeds of the common bean, Phaseolus vulgaris, and PHA-L is theleucocyte-agglutinating form of this lectin. This tetramericglycoprotein accumulates in the vacuoles of storage parenchymacells. Based on amino acid sequence comparisons of legume lectinsand the three-dimensional structure of lectin-carbohydrate complexes,Asn¹²⁸ can be identified as a likely candidate for site-directedmutagenesis to create a mutant PHA-L that does not bind carbohydrate.PHA-L N¹²⁸     

Identification of the G protein-activating sequence of the single-transmembrane natriuretic peptide receptor C (NPR-C)

Rat natriuretic peptideclearance receptor (NPR-C) contains four sequences capable ofinhibiting adenylyl cyclase. We have undertaken mutational and deletionstudies on the intracellular domain of rat NPR-C to determine which ofthese sequences is functionally relevant. Nine mutant receptors wereconstructed by deletion of 11 or 28 COOH-terminal residues or bysite-directed mutagenesis of basic residues in a 17-amino acidsequence, R⁴⁶⁹RNHQEESNIGKHRELR⁴⁸⁵,corresponding to the main active peptide. Substitution of arginine residues (R⁴⁶⁹R⁴⁷⁰) flanking theNH₂ terminus abolished G_i1 and G_i2and PLC- activities and inhibition of adenylyl cyclase. Substitutionof one or two basic residues (H⁴⁸¹ and/or R⁴⁸²or R⁴⁸⁵) in the COOH-terminal motif(H⁴⁸¹RELR⁴⁸⁵) greatly decreased or abolished Gprotein and PLC- activities and inhibition of adenylyl cyclase. Thisimplies that sequences NH₂-terminal to the motif orCOOH-terminal to R⁴⁷⁰ could not sustain receptor activityin situ, although they exhibited activity when used as syntheticpeptides. Deletion of the 11 COOH-terminal residues (E⁴⁸⁶to A⁴⁹⁶) suggested an autoinhibitory function for thissequence. We conclude that the 17-amino acid sequence (R⁴⁶⁹to R⁴⁸⁵) in the middle region of the intracellular domainof NPR-C is both necessary and sufficient for activation of G proteinsand effector enzymes.

     

Tritium labelling of amino sugars at C-2 by alkaline epimerization in tritiated water

N-Acetyl-D-[2-³H]glucosamine was synthesized from N-acetyl-D-mannosamineby alkaline 2-epimerization in pyridine containing ³H₂O andnickelous acetate. The reaction involves reversible formationof an enol intermediate and therefore also resulted in incorporationof tritium into N-acetylmannosamine. After completed reaction,the two N-acetylhexosamines were separated from other radioactiveproducts and Morgan-Elson chromogens by chromatography on acolumn of Sephadex G-10, which was eluted with 10% ethanol,and were then separated from each other by chromatography onSephadex G-15 in 0·27 M sodium borate (pH 7·8).The location of the incorporated tritium was established bytreatment of the N-acetylhexosamines with borate under the conditionsof the Morgan-Elson reaction, which converts the sugars to Kuhn'schromogen I with concomitant loss of the C-2 hydrogen. As expected,this treatment resulted in the formation of ³H₂O, indicatingthat the tritium was located at C-2. [2-³H]Glucosamine was preparedby acid hydrolysis of the labelled N-acetylglucosamine and wasconverted to [2-³H]glucosamine 6-phosphate by incubation withhexokinase and ATP. The sugar phosphate was used as a substratefor glucosamine 6-phosphate deaminase (isomerase, EC 5.3.1.10 [EC] )in a simple ³H₂O release assay. N-acetyl[2-³H]glucosamine N-acetyl[2-³H]mannosamine [2-³H]glucosamine glucosamine 6-phosphate deaminase [2-³H]mannosamine     

黑龙江省落叶松人工林碳储量动态研究

基于36株碳密度测定样木和5期黑龙江省森林资清查数据（1986~2005）,利用非线性度量误差模型来估计黑龙江省落叶松人工林的碳储量动态变化。结果表明：黑龙江省人工落叶松不同器官碳密度在456.7~479.0 mg·g^-1之间,不同器官碳密度差异显著,各器官碳密度由高到低为：树叶>树枝>树干>树根。不同林龄落叶松人工林树干、树根树枝和树叶的碳储量分配比例分别稳定在：66.75%~68.92%、21.59%~22.62%、5.99%~8.16%和2.47%~3.50%。其中,树根和树枝含碳量比重随林分年龄增加而增加,树干和树叶含碳量比重随林分年龄增大而减小。1986~2005年黑龙江省落叶松人工林碳储量总体呈增长趋势,2000年时达最大,为30.38 t·hm^-2,在此期间,平均每年以1.21 t·hm^-2的速度增加。2005年黑龙江省不同区域落叶松人工林碳储量在25.43~34.35 t·hm^-2之间,各区域碳储量由高到低依次为：小兴安岭南坡>完达山地区>张广才岭东坡>张广才岭西坡>小兴安岭北坡。     

Excessive use of nitrogen in Chinese agriculture results in high N2O/(N2O+N2) product ratio of denitrification,primarily due to acidification of the soils

China is the world's largest producer and consumer of fertilizer N, and decades of overuse has caused nitrate leaching and possibly soil acidification. We hypothesized that this would enhance the soils' propensity to emit N₂O from denitrification by reducing the expression of the enzyme N₂O reductase. We investigated this by standardized oxic/anoxic incubations of soils from five long‐term fertilization experiments in different regions of China. After adjusting the nitrate concentration to 2 mM, we measured oxic respiration (R), potential denitrification (D), substrate‐induced denitrification, and the denitrification product stoichiometry (NO, N₂O, N₂). Soils with a history of high fertilizer N levels had high N₂O/(N₂O+N₂) ratios, but only in those field experiments where soil pH had been lowered by N fertilization. By comparing all soils, we found a strong negative correlation between pH and the N₂O/(N₂O+N₂) product ratio (r² = 0.759, P < 0.001). In contrast, the potential denitrification (D) was found to be a linear function of oxic respiration (R), and the ratio D/R was largely unaffected by soil pH. The immediate effect of liming acidified soils was lowered N₂O/(N₂O+N₂) ratios. The results provide evidence that soil pH has a marginal direct effect on potential denitrification, but that it is the master variable controlling the percentage of denitrified N emitted as N₂O. It has been known for long that low pH may result in high N₂O/(N₂O+N₂) product ratios of denitrification, but our documentation of a pervasive pH‐control of this ratio across soil types and management practices is new. The results are in good agreement with new understanding of how pH may interfere with the expression of N₂O reductase. We argue that the management of soil pH should be high on the agenda for mitigating N₂O emissions in the future, particularly for countries where ongoing intensification of plant production is likely to acidify the soils.     

Novel role of the Ca2+-ATPase in NMDA-induced intracellular acidification

The mechanism involved inN-methyl-D-glucamine(NMDA)-induced Ca²⁺-dependentintracellular acidosis is not clear. In this study, we investigated indetail several possible mechanisms using cultured rat cerebellargranule cells and microfluorometry [fura 2-AM or 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein-AM].When 100 µM NMDA or 40 mM KCl was added, a marked increase in theintracellular Ca²⁺ concentration([Ca²⁺]_i)and a decrease in the intracellular pH were seen. Acidosis wascompletely prevented by the use ofCa²⁺-free medium or1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-AM, suggesting that it resulted from an influx of extracellular Ca²⁺. The following fourmechanisms that could conceivably have been involved were excluded:1)Ca²⁺ displacement of intracellularH⁺ from common binding sites;2) activation of an acid loader or inhibition of acid extruders; 3)overproduction of CO₂ or lactate; and 4) collapse of the mitochondrialmembrane potential due to Ca²⁺uptake, resulting in inhibition of cytosolicH⁺ uptake. However,NMDA/KCl-induced acidosis was largely prevented by glycolyticinhibitors (iodoacetate or deoxyglucose in glucose-free medium) or byinhibitors of the Ca²⁺-ATPase(i.e.,Ca²⁺/H⁺exchanger), including La³⁺,orthovanadate, eosin B, or an extracellular pH of 8.5. Our results therefore suggest that Ca²⁺-ATPaseis involved in NMDA-induced intracellular acidosis in granule cells. Wealso provide new evidence that NMDA-evoked intracellular acidosisprobably serves as a negative feedback signal, probably with theacidification itself inhibiting the NMDA-induced[Ca²⁺]_i increase.

     

Optimal alignment between groups of sequences and its application to multiple sequence alignment

Four algorithms, A–D, were developed to align two groupsof biological sequences. Algorithm A is equivalent to the conventionaldynamic programming method widely used for aligning ordinarysequences, whereas algorithms B – D are designed to evaluatethe cost for a deletion/insertion more accurately when internalgaps are present in either or both groups of sequences. Rigorousoptimization of the ‘sum of pairs’ (SP) score isachieved by algorithm D, whose average performance is closeto O(MNL²) where M and N are numbers of sequences included inthe two groups and L is the mean length of the sequences. AlgorithmB uses some app mximations to cope with profile-based operations,whereas algorithm C is a simpler variant of algorithm D. Thesegroup-to-group alignment algorithms were applied to multiplesequence alignment with two iterative strategies: a progressivemethod based on a given binary tree and a randomized grouping-realignmentmethod. The advantages and disadvantages of the four algorithmsare discussed on the basis of the results of exatninations ofseveral protein families.     

Protein Turnover in Isolated Barley Leaf Segments and the Effects of Stress

Leaf segments prepared from the first leaves of barley (Hordeumvulgare) exhibit a rapid loss of protein when given a matricstress with polyethylene glycol. Protein synthesis was reducedby the stress but a greater effect of stress was seen on proteindegradation. Growing leaves were exposed to ³H₂O for 4 d ormore to label total protein, and the half-life of protein 2-³H,in the isolated segments prepared from such leaves, was shownto be c. 140 h in the absence of stress. Stress reduced thisto c. 62 h. A short pulse with ³H₂O preferentially labels rapidlyturning-over protein and a 24 h pulse given to isolated leafsegments labelled proteins with a half-life of c. 64 h in thepresence or absence of stress. Degradation of the 24 h pulse-labelledproteins was inhibited by cycloheximide. Proline accumulationoccurred in the stressed segments and was inhibited by cycloheximide.The results are discussed in the light of current views concerningprotein degradation and possible relationships between proteolysisand proline accumulation.     

Variations in the Contents and Kinetic Properties of Ribulose-1,5-bisphosphate Carboxylases among Rice Species

The K_m(CO₂) ancl V_max of ribulose 1,5-bisphosphate (RuBP) carboxylaseand its protein ratio to total soluble protein from Oryza speciesincluding cultivars (25 varieties) and wild types (11 species,21 strains) were surveyed. Their variabilities among cultivarsof O. sativa were very small. The averages of the K_m(CO₂) andV_max values and the ratio of carboxylase to soluble protein,and their standard errors were 10.2?1.0µM, 1.72?0.13units.mg^–1(pH 8.0 and 25?C) and 52?2%, respectively. However, some differencesseemed to exist based on genome constitution in the Oryza genus.RuBP carboxylases from the species with the A^gA^g genome, O.graberrima and O. breviligulate, exhibited low K_m(CO₂) values(8.0?0.8 µM). High V_max was associated with the CC genome,O. eichingeri and O. officinalis (2.08?0.15 units.mg^–1).A higher ratio of RuBP carboxylase protein to soluble proteinwas found for the AA genome, O. sativa and O. perennis. (Received September 24, 1986; Accepted April 15, 1987)     

鼎湖山主要森林土壤N2O排放及其对模拟N沉降的响应

研究了鼎湖山生物圈保护区马尾松(Pinus massoniana)林、混交林和季风常绿阔叶林(季风林)土壤N₂O排放特征及其对氮沉降增加的响应。在1999~2002年期间,3种森林土壤N₂O排放速率均表现明显的季节性变化特点,但这种季节性变化因年份和森林类型不同而异,总的来说,3种森林土壤N₂O排放速率呈现夏秋季较高而冬春季较低的变化。土壤N₂O排放速率在3年观测期间的平均值分别为(g·hm^-2·d^-1):14.2±3.1(季风林),5.8±0.9(混交林)和5.1±0.9(马尾松林)。土壤N₂O排放速率与土壤温度之间在季风林呈现显著的指数正相关关系,但在混交林和马尾松林中它们之间的关系则均不明显。经3个月的模拟氮沉降试验后,氮沉降增加对季风林和马尾松林土壤N₂O的排放均具有明显的促进作用,且这种促进作用随氮沉降水平的升高而增强,但对混交林土壤N₂O排放的影响则不明显。     

Approximate matching of regular expressions

Given a sequenceA and regular expressionR, theapproximate regular expression matching problem is to find a sequence matchingR whose optimal alignment withA is the highest scoring of all such sequences. This paper develops an algorithm to solve the problem in timeO(MN), whereM andN are the lengths ofA andR. Thus, the time requirement is asymptotically no worse than for the simpler problem of aligning two fixed sequences. Our method is superior to an earlier algorithm by Wagner and Seiferas in several ways. First, it treats real-valued costs, in addition to integer costs, with no loss of asymptotic efficiency. Second, it requires onlyO(N) space to deliver just the score of the best alignment. Finally, its structure permits implementation techniques that make it extremely fast in practice. We extend the method to accommodate gap penalties, as required for typical applications in molecular biology, and further refine it to search for substrings ofA that strongly align with a sequence inR, as required for typical data base searches. We also show how to deliver an optimal alignment betweenA andR in onlyO(N+logM) space usingO(MN logM) time. Finally, anO(MN(M+N)+N ²logN) time algorithm is presented for alignment scoring schemes where the cost of a gap is an arbitrary increasing function of its length.