Gene product of v-fgr onc: hybrid protein containing a portion of actin and a tyrosine-specific protein kinase

The nucleotide sequence of the region of Gardner-Rasheed feline sarcoma virus (GR-FeSV) encoding its primary translation product, p70gag-fgr, has been determined. From the nucleotide sequence, the amino acid sequence of this transforming protein was deduced. Computer analysis indicates that a portion of P70gag-fgr has extensive amino acid sequence homology with actin, a eukaryotic cytoskeletal protein. A second region of P70gag-fgr is closely related to the tyrosine-specific kinase gene family. Thus, the v-fgr oncogene appears to have arisen as a result of recombinational events involving two distinct cellular genes, one coding for a structural protein and the other for a protein kinase.     

Primary structure of v-raf: relatedness to the src family of oncogenes

A replication-defective, acute transforming retrovirus (murine sarcoma virus 3611) was isolated from mouse and molecularly cloned. The nucleotide sequence of 1.5 kilobases encompassing the transforming gene (v-raf) was determined. This sequence, which predicts the amino acid sequence of a gag-raf fusion protein, terminates 180 nucleotides from the 3' end of the acquired cellular sequence. Comparison of the predicted amino acid sequence of v-raf with the predicted amino acid sequences of other oncogenes reveals significant homologies to the src family of oncogenes. There is a lack of homology within the sequence of the tyrosine acceptor domain described for the phosphotyrosine kinase members of the src family of transforming proteins. Phylogenetic arrangement of this family of oncogenes suggests that tyrosine-specific phosphorylation may be a recently acquired activity.     

The E5 transforming gene of bovine papillomavirus encodes a small, hydrophobic polypeptide

Bovine papillomavirus (BPV-1) contains two independent transforming genes that have been mapped to the E5 and E6 open reading frames (ORF's). The E5 transforming protein was identified by means of an antiserum against a synthetic peptide corresponding to the 20 COOH-terminal amino acids of the E5 ORF. The E5 polypeptide is the smallest viral transforming protein yet characterized; it had an apparent size of 7 kilodaltons. The transforming polypeptide is encoded entirely within the second half of the E5 ORF and its predicted amino acid composition is very unusual; 68% of the amino acids are strongly hydrophobic and 34% are leucine. Cell fractionation studies localized this polypeptide predominantly to cellular membranes.     

Expression of a platelet-derived growth factor-like protein in simian sarcoma virus transformed cells

The near identity of the partial amino acid sequence of human platelet-derived growth factor (PDGF) and that predicted for p28sis, the putative transforming protein of the simian sarcoma virus (SSV), suggests expression of a growth factor activity may be central for transformation by SSV. It is now reported that SSV-transformed cells but not control cells contain a growth factor activity that is identical to PDGF in immunoassay, in mitogenic dose response, and in specific mitogenic activity. The protein immunoprecipitated by antiserum to human PDGF has an apparent molecular weight of 20,000, identical to that of p20sis, the putative intracellular degradation product of p28sis. The results support the concept that expression of a PDGF-like molecule, which appears to be the product of the viral-sis gene, is responsible for the abnormal regulation of growth is SSV-transformed cells.     

Three-dimensional structure of an oncogene protein: catalytic domain of human c-H-ras p21

The crystal structure at 2.7 A resolution of the normal human c-H-ras oncogene protein lacking a flexible carboxyl-terminal 18 residue reveals that the protein consists of a six-stranded beta sheet, four alpha helices, and nine connecting loops. Four loops are involved in interactions with bound guanosine diphosphate: one with the phosphates, another with the ribose, and two with the guanine base. Most of the transforming proteins (in vivo and in vitro) have single amino acid substitutions at one of a few key positions in three of these four loops plus one additional loop. The biological functions of the remaining five loops and other exposed regions are at present unknown. However, one loop corresponds to the binding site for a neutralizing monoclonal antibody and another to a putative "effector region"; mutations in the latter region do not alter guanine nucleotide binding or guanosine triphosphatase activity but they do reduce the transforming activity of activated proteins. The data provide a structural basis for understanding the known biochemical properties of normal as well as activated ras oncogene proteins and indicate additional regions in the molecule that may possibly participate in other cellular functions.     

多肽和蛋白质的人工合成评述

多肽和蛋白质的人工合成,就是多个氨基酸在一定的条件下通过肽键形成多肽链。氨基酸是合成多肽和蛋白质的原料。在合成前各个氨基酸都必须经过基团保护、氨基和羧基的活化处理,然后经接肽,去除保护基,最后形成多肽链。本文对目前国内外常用的液相合成法和固相合成法的原理和步骤进行了简要的评述。     

普通荞麦种内丝裂原活化蛋白激酶序列分析

【目的】比较普通荞麦Fagopyrum esculentum种内丝裂原活化蛋白激酶基因(MAPK)序列的差异,研究MAPK基因序列在普通荞麦栽培进化过程中的变化。【方法】以普通荞麦的9个栽培品种和3个落花落果野生种质为材料,PCR特异性扩增获得MAPK基因的保守片段,对基因片段序列进行差异分析和蛋白结构预测。【结果】荞麦MAPK基因cDNA全长为2 835 bp,开放阅读框1 827 bp,编码609个氨基酸,含有TDY的三肽模块,为植物D组MAPK蛋白。PCR扩增获得12个供试材料的MAPK序列,其单型不变位点为723个,多态位点为70个。9个栽培品种间开放阅读框(ORF)区域无序列差异,3个野生种质间ORF区域也无序列差异。栽培品种与野生品种的ORF区域序列含有8个差异位点,编码3个差异氨基酸。其中,ORF区域第13位点组氨酸(H)→酪氨酸(Y)发生置换,导致1个α-螺旋构象发生变化。【结论】普通荞麦MAPK基因序列高度保守,栽培驯化对ORF区域第13位点差异氨基酸的选择具有高度一致性。     

荞麦芽菜蛋白质营养的评价研究

对甜荞 (Fagopyrum esculentum)和苦荞 (F.tartaricum)芽的蛋白质营养进行分析 ,并应用氨基酸比值系数法 ,对荞麦芽的氨基酸进行全面评价。结果表明 ,苦荞 (榆 6- 2 1 )芽的粗蛋白含量为 1 42 .91 g/kg,甜荞 (榆荞 1号 )芽的粗蛋白含量为 1 72 .72 g/kg。二者蛋白质中氨基酸种类齐全 ,必需氨基酸 (EAA)分别占总氨基酸的 43.1 9%和 40 .89%。第一限制性氨基酸为含硫氨基酸 (Met+Cys)。甜荞芽菜的氨基酸比值系数分为 81 .97,略高于鸡蛋 ,苦荞芽菜的氨基酸比值系数分为 74.45,其蛋白质营养优于一般的蔬菜。抗营养因子检测未发现胰蛋白酶抑制剂活性。     

玉米籽粒蛋白质构成的多元回归与聚类分析

以49个普通玉米杂交组合为材料,分析了玉米籽粒蛋白质及其构成的16种氨基酸含量间的多元相关与回归关系。结果表明,大多数氨基酸及其与蛋白质含量之间存在显著的正相关关系;各氨基酸与蛋白质含量之间分别有4种呈线性回归关系,5种呈二次曲线关系,7种呈三次曲线关系。以营养品质性状间的皮尔逊相关系数做测度指标,采用系统聚类的方法可将蛋白质及16种氨基酸含量聚为4个大类。对各类中的典型性状如赖氨酸、丝氨酸、丙氨酸和天冬氨酸含量的选择,可能会有效地提高玉米品质育种中蛋白质构成的改良效率。     

Mechanisms of antibody binding to a protein

The mechanisms of antibody binding to a protein were studied by an analysis of specific amino acid residues critical to nine antigenic sites on myohemerythrin. Rabbit antisera to the whole protein were assayed for binding to more than 1500 distinct peptide analogs differing from the protein sequence by single amino acid replacements. The results, combined with information from the three-dimensional crystallographic structure, were used to evaluate probable mechanisms of antibody binding at individual sites. The data from all sites examined indicate that initial binding to solvent-exposed amino acid residues may promote local side-chain displacements and thereby allow the participation of other, previously buried, residues.     

Microtubule-associated protein MAP2 shares a microtubule binding motif with tau protein

The microtubule-associated protein MAP2 is a prominent large-sized component of purified brain microtubules that, like the 36- to 38-kilodalton tau proteins, bears antigenic determinants found in association with the neurofibrillary tangles of Alzheimer's disease. The complete sequence of mouse brain MAP2 was determined from a series of overlapping cloned complementary DNAs. The sequence of the carboxyl-terminal 185 amino acids is very similar (67 percent) to a corresponding region of tau protein, and includes a series of three imperfect repeats, each 18 amino acids long and separated by 13 or 14 amino acids. A subcloned fragment spanning the first two of the 18-amino acid repeats was expressed as a polypeptide by translation in vitro. This polypeptide copurified with microtubules through two successive cycles of polymerization and depolymerization, whereas a control polypeptide derived from the amino-terminal region of MAP2 completely failed to copurify. These data imply that the carboxyl-terminal domain containing the 18-amino acid repeats constitutes the microtubule binding site in MAP2. The occurrence of these repeats in tau protein suggests that these may be a general feature of microtubule binding proteins.     

DTA-6对紫花苜蓿粗蛋白和氨基酸含量的调控作用

以敖汉苜蓿(Medicago sativa)为材料研究了新型植物生长调节剂DTA-6对苜蓿叶片、茎杆中粗蛋白、氨基酸、必需氨基酸和含硫氨基酸含量的影响。研究表明DNA-6处理后苜蓿叶片粗蛋白含量比对照增加了39．2％，茎杆中粗蛋白比对照增加了33．5％。DNA-6处理提高了叶片中含硫氨基酸和必需氨基酸总量，降低了茎杆中含硫氨基酸和必需氨基酸总量，而且调节作用的剂量效应明显。对DNA-6的作用机理作了初步探讨。     

芜菁花叶病毒杭州分离株外壳蛋白基因序列分析

从杭州郊区分离出一个ＴｕＭＶ强毒株系，利用分子克隆和Ｓａｎｇｅｒ双脱氢测序方法分析了其外壳蛋白基因的ＤＮＡ序列．该序列与已报道的其他４种ＴｕＭＶ分离物都具有较高的同源性，最高达９７．６％，最低达８９．５％，其氨基酸序列的同源性更高，最高达９７．９％，最低达９４．５％。本文分析了该序列中ＡＴ和ＧＣ含量，计算了４种核苷酸在有意义链中和在密码子不同位置上的出现频率，同时还分析了该病毒外壳蛋白氨基酸遗传密码的使用频率。     

提高豆科植物蛋氨酸含量的基因工程途径

豆科植物普遍缺乏蛋氨酸和半胱氨酸这类含硫氨基酸,影响了其营养品质。采用基因工程手段改造内源蛋白基因,克隆异源高蛋氨酸蛋白基因,人工合成氨基酸平衡的蛋白基因,以及改造或过表达氨基酸代谢通路中的关键酶基因,然后通过转基因手段转化豆科植物,会使蛋氨酸含量均有不同程度提高。另外,利用高蛋氨酸的近缘种质资源进行有性杂交育种,通过化学诱变培育高蛋氨酸突变株,也可获得一些高蛋氨酸新品系。通过对这些方法的比较分析,提出了提高蛋氨酸丰富蛋白表达量和蛋氨酸合成量两方面相结合的研究思路,为豆科植物品质改良研究提供参考。     

Encephalitogenic protein: structure

Amino acid sequences of encephalitogenic proteins from bovine cord and rabbit brain are reported. The bovine protein contains 45 residues. The rabbit protein is identical except for two isopolar substitutions, a dipeptide and amino acid deletion. Analysis of this protein and a 140-residue myelin basic protein indicates that the smaller protein is a portion of the larger encephalitogen. The larger myelin protein contains at least two encephalitogenic sites.     

The oncogenic activation of human p21ras by a novel mechanism

Single amino acid changes were introduced into normal (non-oncogenic) and activated forms of the human H-ras protein at a position (residue 116) proposed on structural grounds to represent a contact site with guanine nucleotides. Substitutions at this site could significantly reduce the ability of both forms to bind and hydrolyze guanosine 5'-triphosphate; these substitutions, however, did not necessarily diminish the transforming capacity of activated derivatives. One substitution that severely impairs these functions activated the transforming potential of the otherwise normal polypeptide.     

In vivo half-life of a protein is a function of its amino-terminal residue

When a chimeric gene encoding a ubiquitin-beta-galactosidase fusion protein is expressed in the yeast Saccharomyces cerevisiae, ubiquitin is cleaved off the nascent fusion protein, yielding a deubiquitinated beta-galactosidase (beta gal). With one exception, this cleavage takes place regardless of the nature of the amino acid residue of beta gal at the ubiquitin-beta gal junction, thereby making it possible to expose different residues at the amino-termini of the otherwise identical beta gal proteins. The beta gal proteins thus designed have strikingly different half-lives in vivo, from more than 20 hours to less than 3 minutes, depending on the nature of the amino acid at the amino-terminus of beta gal. The set of individual amino acids can thus be ordered with respect to the half-lives that they confer on beta gal when present at its amino-terminus (the "N-end rule"). The currently known amino-terminal residues in long-lived, noncompartmentalized intracellular proteins from both prokaryotes and eukaryotes belong exclusively to the stabilizing class as predicted by the N-end rule. The function of the previously described posttranslational addition of single amino acids to protein amino-termini may also be accounted for by the N-end rule. Thus the recognition of an amino-terminal residue in a protein may mediate both the metabolic stability of the protein and the potential for regulation of its stability.     

Rapid and sensitive protein similarity searches

An algorithm was developed which facilitates the search for similarities between newly determined amino acid sequences and sequences already available in databases. Because of the algorithm's efficiency on many microcomputers, sensitive protein database searches may now become a routine procedure for molecular biologists. The method efficiently identifies regions of similar sequence and then scores the aligned identical and differing residues in those regions by means of an amino acid replacability matrix. This matrix increases sensitivity by giving high scores to those amino acid replacements which occur frequently in evolution. The algorithm has been implemented in a computer program designed to search protein databases very rapidly. For example, comparison of a 200-amino-acid sequence to the 500,000 residues in the National Biomedical Research Foundation library would take less than 2 minutes on a minicomputer, and less than 10 minutes on a microcomputer (IBM PC).     

Visualization of amino acid composition differences between processed protein from different animal species by self-organizing feature maps

Amino acids are the dominant organic components of processed animal proteins, however there has been limited investigation of differences in their composition between various protein sources. Information on these differences will not only be helpful for their further utilization but also provide fundamental information for developing species-specific identification methods. In this study, self-organizing feature maps (SOFM) were used to visualize amino acid composition of fish meal, and meat and bone meal (MBM) produced from poultry, ruminants and swine. SOFM display the similarities and differences in amino acid composition between protein sources and effectively improve data transparency. Amino acid composition was shown to be useful for distinguishing fish meal from MBM due to their large concentration differences between glycine, lysine and proline. However, the amino acid composition of the three MBMs was quite similar. The SOFM results were consistent with those obtained by analysis of variance and principal component analysis but more straightforward. SOFM was shown to have a robust sample linkage capacity and to be able to act as a powerful means to link different sample for further data mining.