Amino acid sequence of the fetal chain of yak haemoglobin

The amino acid sequence of the fetal chain of yak haemoglobin was determined. The sequence is the same as that of the fetal chain of bovine haemoglobin. Phenylalanine is present at position 12 of the helix A in the fetal chain while tryptophan is the amino acid at this position in theβ-chain of yak adult haemoglobin. This amino acid replacement may be responsible for the higher oxygen affinity of yak fetal haemoglobin than yak adult haemoglobin.     

Studies of haemoglobin types in barbary sheep (Ammotragus lervia)

1. Two haemoglobin types, haemoglobins Amm-C and Amm-B, were observed in five Barbary sheep (Ammotragus lervia). One animal was homozygous for haemoglobin Amm-C, a second was homozygous for haemoglobin Amm-B, and three were heterozygous for both. 2. Amino acid analyses of the globin from haemoglobin Amm-B showed that this type was related to, but not identical with, haemoglobin B of the domestic sheep. 3. The β-chain of haemoglobin Amm-C was found to be composed of 141 amino acid residues. Its amino acid composition differed from that of the β^C-chain of the anaemic domestic sheep in at least 14 residues. The Amm-β^C-chain contained one isoleucyl residue. 4. The amino acid compositions of tryptic peptides T-1, T-2, T-13 and T-14 of the Amm-β^C-chain were similar to those of the sheep β^C-chain. Peptides T-3, T-4, T-6, T-7, T-8, T-11 and T-15 were the same as the corresponding peptides of the sheep β^A- and β^C-chains. Peptide T-5 and to a smaller extent peptide T-9 resembled the corresponding peptides of the sheep β^A-chain, and peptide T-10 was identical with peptide γT-10 of sheep haemoglobin F. Peptide T-12 was not recovered. 5. The results of these investigations were interpreted as being indicative that the structural Amm-β^C-gene is closely related to the β^C-gene of sheep, from which through domestication the present domestic sheep originated.     

Haemoglobin I: a new β-globin chain variant found in sheep of Italian breeds

A rather common haemoglobin variant was detected in the Sardinian and Altamurana sheep breeds. The mutated globin chain appears to be produced under the control of an allele at the HBB locus and due to a neutral amino acid substitution. The variant will be provisionally referred to as the Hb I.     

A comparison of the amino acid sequences of Crithidia fasciculata and Trypanosoma rhodesiense cytochromes c

Apocytochrome c was isolated from procyclic trypomastigotes of Trypanosoma rhodesiense EATRO 1895 and purified on Amberlite IRC-50 ion-exchange resin. Tryptic peptides were generated from the purified apoprotein and a partial amino acid sequence was determined. A comparison of the amino acid sequence of Crithidia fasciculata with the partial amino acid sequence of T. rhodesiense reveals significant homology.     

A cereal haemoglobin gene is expressed in seed and root tissues under anaerobic conditions

Legumes, and a very few non-legume plant species, are known to possess functioning haemoglobin genes. We describe here the characterization of a haemoglobin cDNA isolated from barley. The deduced amino acid sequence shows 71% amino acid identity with a non-legume haemoglobin gene, a further 16% of the residues being conservative replacements. The barley cDNA also hybridizes to genomic sequences in rye, maize and wheat. The demonstration of a gene from a monocotyledon with close sequence homology to the known non-legume plant haemoglobins fills a major gap in the known distribution of haemoglobin genes in the plant kingdom. The expression of the gene is induced in isolated barley aleurone layers exposed to anaerobic conditions, and the roots of flooding-stressed barley plants. The expression of the RNA under anoxic conditions is similar to that of a known anaerobic response gene, alcohol dehydrogenase. Our results suggest that the increased expression of haemoglobin RNA is an integral part of the normal anaerobic response in barley. The findings are discussed in the light of current theories of haemoglobin function and evolution.     

Sequence analysis of UTR and coding region of kappa-casein gene of Indian riverine buffalo (Bubalus bubalis).

In this study, complete nucleotide as well as derived amino acid sequence characterization of water buffalo (Bubalus bubalis) kappa-casein gene has been presented. Kappa-casein cDNA clones were identified and isolated from a buffalo lactating mammary gland cDNA library. Sequence analysis of kappa-casein cDNA revealed 850 nucleotides with an open reading frame (ORF) of 573 nucleotides, encoding mature peptide of 169 amino acids. The 5' untranslated region (UTR) comprised 71 nucleotides, while 3' UTR was of 206 nucleotides. A total of 11 nucleotide and seven amino acid changes were observed in, buffalo (Bubalus bubalis) as compared to cattle (Bos taurus), sheep (Ovis aries) and goat (Capra hircus). Among these nucleotide changes, eight were unique in buffalo as they were fully conserved in cattle, sheep and goat. Majority of the nucleotide changes and all the amino acid changes; 14 (Asp-Glu), 19(Asp/Ser-Asn), 96(Ala-Thr), 126(Ala-Val), 128(Ala/Gly-Val), 156(Ala/Pro-Val) and 168(Ala/Glu-Val) were limited to exon IV. Three glycosylation sites, Thr 131, Thr 133 and Thr 142 reported in cattle and goat kappa-casein gene were also conserved in buffalo, however, in sheep Thr 142 was replaced by Ala. Chymosin hydrolysis site, between amino acids Phe 105 and Met 106, important for rennet coagulation process, were found to be conserved across four bovid species. Buffalo kappa-casein with the presence of amino acids Thr 136 and Ala 148 seems to be an intermediate of "A" and "B" variants of cattle. Comparison with other livestock species revealed buffalo kappa-casein sharing maximum nucleotide (95.5%) and amino acid (92.6%) similarity with cattle, whereas with pig it showed least sequence similarity of 76.0% and 53.2%, respectively. Phylogenetic analysis based on both nucleotide and amino acid sequence indicated buffalo kappa-casein grouping with cattle, while sheep and goat forming a separate cluster close to them. The non-ruminant species viz. camel, horse and pig were distantly placed, in separate lineages.     

Sheep haemoglobin i or β b13(a10)gly → ser: an example of a cpg mutation in vertebrates. Characterization using fab-mass spectrometry and amino acid sequencing

• 1.1. The amino acid substitution which characterizes the haemoglobin I variant from sheep has been ascertained using a combination of Fast Atom Bombardment mass spectrometry and protein sequencing.
• 2.2. A Ser for Gly substitution at position 13 (10 of the A helix) was found in a polypeptide with the overall sequence of the β^B globin.
• 3.3. On the basis of the nucleotide sequence of the β^B-globin gene, a C to T transition occurring on a CpG doublet is considered to be responsible for the amino acid substitution.
• 4.4. This represents the first observation of a variant sheep Hb due to a mutation which is rather common in the human genome.
• 5.5. Amongst ruminants, serine is normally present at position 13 of goat and sheep ε¹¹ and γ chains and of bovine γ chain which had an independent and more ancient evolutionary origin than the β chains.

     

Isolation and characterization of a complementary DNA clone coding for the E1 beta subunit of the bovine branched-chain alpha-ketoacid dehydrogenase complex: complete amino acid sequence of the precursor protein and its proteolytic processing

A 1.7-kb cDNA clone encoding the entire precursor of the E1 beta subunit of the branched-chain alpha-ketoacid dehydrogenase (BCKDH) complex was isolated from a bovine liver cDNA library by screening with a mixture of synthetic oligonucleotide probes corresponding to the C-terminal five-residue sequence of the mature E1 beta subunit. A partial amino acid sequence was determined by Edman degradation of the intact subunit and the peptides generated by cleavage at the lysyl bonds. Nucleotide sequence analysis revealed that the isolated cDNA clone contained the 5'-untranslated sequence of 186 nucleotides, the translated sequence of 1176 nucleotides, and the 3'-untranslated sequence of 306 nucleotides with a poly(A) tail. A type AATAAA polyadenylation signal was located 17 nucleotides upstream of the start of a poly(A) tail. Comparison of the amino acid sequence predicted from the nucleotide sequence of the cDNA insert of the clone with the partial amino acid sequence of the mature BCKDH E1 beta subunit showed that the cDNA insert encodes for a 342 amino acid subunit with Mr 37,745 and that the subunit is synthesized as the precursor with a leader sequence of 50 amino acids and processed at the N-terminus. Northern blot analysis using the cDNA insert as a probe showed the presence of a 1.8-1.9-kb mRNA in bovine liver, suggesting that the insert covers nearly a full length of mRNA. Alignment of the deduced amino acid sequence of bovine BCKDH E1 beta with that of the human pyruvate dehydrogenase (PDH) complex E1 beta subunit revealed a high degree of sequence homology throughout the two enzymes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Subunit assembly of giant haemoglobin from the polychaete Tylorrhynchus heterochaetus

The subunit assembly of the giant haemoglobin of the polychaete Tylorrhynchus heterochaetus is presented. Tylorrhynchus haemoglobin consists of two types of subunits: a "monomeric" chain I and a disulphide-bonded "trimer" of chains IIA, IIB and IIC. The molar ratio of the four constituent chains was determined by statistical comparison of the accurate amino acid composition calculated from the sequence of each chain and the observed composition measured by amino acid analysis of the whole molecule. On the basis of the molar ratio and the molecular weight of each chain, deduced from the amino acid sequence, a symmetrical model for the molecular assembly of the haemoglobin was constructed. The proposed model consists of four species of chains of 192 polypeptides and has a molecular weight of 3,275,808. The minimum structural entity is a "tetramer" consisting of the "monomeric" chain and the disulphide-bonded "trimer". Each chain contains one haem.     

绵羊肺腺瘤病毒中国NM株部分gag基因的克隆与序列分析

参照GenBank中已发表的绵羊肺腺瘤病毒的基因序列,设计合成一对引物,对绵羊肺腺瘤病毒(Jaagsiekte sheep retrovirus,JSRV)内蒙株的gag基因中主要编码CA蛋白的基因段进行PCR扩增,产物经琼脂糖凝胶电泳分析,呈现一条约897bp的特异条带,将其回收后克隆入pMD—18T载体中,并进行序列测定与分析。结果表明,与南非株(基因序列号NC—001494)的gag基囚序列比较,核苷酸同源性为83％,推导出的氨基酸同源性为84％。与美国株(基因序列号AF105220)的gag基因序列比较,核苷酸同源性为81．5％,氨基酸同源性为83％。这是我国首次报道的绵羊肺腺瘤病毒的gag基因的一段序列,为我国科研工作者进行更深入的研究奠定基础。     

Primary structures of two types of alpha-subunit of rat brain Na+,K+,-ATPase deduced from cDNA sequences

A rat brain cDNA library was screened by using as a probe a fragment of cDNA encoding the alpha-subunit of human Na+,K+-ATPase. Two different cDNA clones were obtained and analyzed. One of them was concluded to be a cDNA encoding the alpha-subunit of the weakly ouabain-sensitive rat kidney-type Na+,K+-ATPase. The deduced amino acid sequence consists of 1,018 amino acids. The alpha-subunit of the rat kidney-type Na+,K+-ATPase shows 97% homology in amino acid sequence with the alpha-subunit of human, sheep, or pig enzyme and 87% with that of Torpedo. Based on a comparison of the amino acid sequence at the extracellular domain of the alpha-subunit between weakly ouabain-sensitive rat kidney-type enzyme and the ouabain-sensitive human, sheep, pig, or Torpedo enzyme, it was proposed that only two significant amino acid replacements are unique to the rat kidney-type alpha-subunit. Another cDNA clone obtained showed 72% homology in nucleotide sequence with the former cDNA coding the alpha-subunit of the rat kidney-type Na+,K+-ATPase and the deduced amino acid sequence exhibited 85% homology with that of the alpha-subunit of rat kidney-type Na+,K+-ATPase.     

Molecular cloning and polymorphism analysis of the prion protein gene in Tan sheep of Ningxia, China

The resistance or susceptibility of sheep to scrapie is associated with polymorphisms of the prion protein gene (PRNP), particularly, single nucleotide polymorphisms (SNPs) in amino acid positions 136, 154 and 171. The prion protein (PrP) gene sequence and the deduced amino acid alignment of prion protein in Tan sheep, a local Chinese sheep breed traditionally raised in Ningxia, northwestern China, were determined and variability of the PrP amino acids sequence was analyzed in this study. The PrP nucleic acids and amino acids sequences of 112 Tan sheep were highly homogenous, although polymorphism of the PrP gene was detected at several sites, particularly codons 106, 154, and 171. The analysis of both sequences revealed that the most predominant allele at codons 136, 154 and 171 in Tan sheep was ARQ, which was known to be associated with high susceptibility to scrapie in sheep. The result suggests that Tan sheep is potentially susceptible to scrapie. Our findings provide valuable information for future breeding projects to scrapie resistance in Tan sheep.     

Primary structure of cyanogen bromide fragments of sei whale (Balaenoptera borealis) pituitary somatotropin]

5 fragments are isolated after the degradation of somatotropin from sei whale pituitary glands with cyanogen bromide: N-terminal 4-segmented; C-terminal 12-segmented with the internal disulfide bond; middle 25- and 30-segmented and a high molecular weight fragment following N-terminal tetrapeptide and bound with disulfide bond to 30-segmented fragment. Complete amino acid sequence of three shortest cyanogen bromide fragments is deciphered and N- and C-terminal sequence is investigated in two large fragments after their uncoupling under performic acid oxidation. Amino acid sequence is deciphered of a peptide obtained after trypsine hydrolysis of 30-segmented cyanogen bromide fragment. Comparison of amino acid sequence of whale somatotropin fragments with that of sheep, beef and human somatotropin has revealed that 57 out of 61 identified amino acid residues of whale somatotropin repeat amino acid residues in similar regions of beef somatotropin, 56--of sheep and only 42--of human somatotropins. Besdies, 4 of 5 revealed amino acid substitutions in whale hormone, as compared with sheep somatotropin, are amino acids which are present at the same positions in human hormone.     

Amino acid sequence of the beta-chain of the tetrameric haemoglobin of the bivalve mollusc, Anadara trapezia

The amino acid sequence of the beta-chain of the principal haemoglobin from A. trapezia has been determined. The sequence was deduced from the sequences of tryptic peptides, which were fractionated using highperformance liquid chromatography and peptide mapping. Additional sequence data, particularly for the large tryptic peptides, was obtained from enzyme digests of both cyanogen bromide fragments and large citraconyltryptic peptides. The beta-chain has 151 residues which is longer than all the other sequenced haemoglobin chains except the alpha-chain of A. trapezia, which is 153 residues in length. The residues corresponding to those normally in the D helix are absent in this beta-chain. The additional residues are contributed by an extension of the N-terminal region, which was also found to be acetylated. Comparison of the beta-chain amino acid sequence with that of the alpha-chain of A. trapezia, the dimeric chain of A. trapezia, and the dimeric chain of A. broughtonii showed 53% identity in each case. In the E and F helices, the homology is particularly noticeable. There is 100% homology in the F helix of all four chains. The dimeric globin of A. trapezia also shows 100% homology with the beta-chain in the E helix, while the alpha-chain shows 75%. If the tertiary structure of the alpha- and beta-chains of A. trapezia haemoglobin is the same as that of horse haemoglobin, then there are many changes in the alpha 1 and beta 2 contact site residues.     

Rat gastric prepepsinogen: in vitro synthesis and partial amino-terminal signal sequence

The major translation product of rat gastric mucosa RNA in a wheat germ cell-free system was identified as prepepsinogen by electrophoretic analysis of its immunoprecipitate on sodium dodecyl sulfate (SDS)-polyacrylamide gels and amino-terminal sequence determination. The translation product containing radioactive amino acids, purified by SDS-polyacrylamide gel electrophoresis, was shown to have an amino-terminal extension peptide comprising 16 amino acid residues. A partial amino acid sequence of this extension peptide is as follows: Met-X-X-Met-Val-Val-X-Leu-Leu-X-Leu-X-Leu-Leu-X-X-pepsinogen.     

The amino acid sequence of myoglobin from skeletal muscles of red deer(Cervus elaphus)

Red deer myoglobin has been fragmented by restricted tryptic digestion and by treatment with cyanogen bromide. The fragments have been separated by gel permeation. The core peptide derived from cyanogen bromide cleavage have been further digested with trypsin and the resulting peptides have been separated on Dowex 1X2. All fragments have been characterized by their amino acid composition, by determination of their N-terminal sequence using automatic Edman degradation and of their C-terminal sequence following the kinetics of amino acid cleavage by carboxypeptidases A and B. The complete sequence has been found to be identical with the already known sequence of sheep myoglobin except for residue 145 which is Gln in red deer globin and Glu in sheep globin. Reinvestigation of the corresponding sequence in sheep globin has shown that residue 145 of sheep globin is also Gln.     

青藏高原藏羚羊朊蛋白基因及氨基酸序列分析

羊瘙痒病是一种自然发生的传染性海绵状脑病,它可以在绵羊和山羊羊群之中传播,其疾病的易感性、潜伏期和跨物种传播的能力主要受宿主朊蛋白基因(prion protein gene,PRNP)的影响。研究藏羚羊PRNP有助于明确藏羚羊和羊瘙痒病之间的关系,从而更好地保护藏羚羊。参照GenBank上已经发表的绵羊PRNP序列设计引物,然后对藏羚羊的PRNP序列进行扩增、测序和分析。测序结果显示藏羚羊PRNP序列由771个核苷酸构成,编码256个氨基酸。序列分析结果显示21只藏羚羊PrP(prion protein,PrP)氨基酸序列完全同源,并且与野生型绵羊PrP氨基酸序列完全一致。此外,藏羚羊PrP氨基酸序列与山羊(99.2%)、汤氏瞪羚(99.2%)、印度羚(99.2%)、驯鹿(98%)、马鹿(98%)、家牛(97.7%)和水牛(96.1%)也具有较高的同源性。在对PrP氨基酸序列136、154、171位多态性分析后发现本次收集的21只藏羚羊样本可能和野生型绵羊一样对羊瘙痒病易感。本研究为羊瘙痒病在藏羚羊羊群中可能出现的风险提供了理论依据,提示要加强藏羚羊羊群中羊瘙痒病的监测。     

Sequence analysis and tissue expression of a non-Bohr beta-globin cDNA from Atlantic salmon

The presence of a haemoglobin protein which does not exhibit a Bohr effect has been found only in fish living in fast flowing waters. We report the cloning of the first non-Bohr effect beta-globin cDNA from an adult Atlantic salmon kidney bank. Nucleotide sequence analysis of this cDNA shows that the predicted beta-globin peptide comprises 147 amino acids with a calculated molecular mass of 15 975 Da and an overall amino acid homology of 40 to 50% to higher vertebrates and 60–90% to fish sequences. This sequence confirms the important amino acid residues which are changed thus causing loss of the Bohr effect [Powers, D.A. and Edmunson, A.B. (1972) Multiple hemoglobins of catostomid fish. J. Biol. Chem. 247, 6686–6693; Brunori, M. (1975) Molecular adaptation to physiological requirements: the hemoglobin system of trout. Curr. Topics Cell. Regul. 9, 1–39]. This loss allows the haemoglobin protein to have a higher oxygen affinity, as it does not release oxygen when the pH of the surrounding environment decreases, which is an important ability for the fish in times of stress.     

Structural characterization of a murine myeloid leukaemia inhibitory factor

A leukaemia inhibitory factor (LIF) which induces macrophage differentiation in M1 murine myeloid leukaemia cells and suppresses their proliferation in vitro has been isolated in sufficient quantities (30 micrograms) from Krebs ascites tumour cell conditioned medium to permit its partial characterization by amino acid sequence analysis. The combination of sensitive microbore column (1.0 and 2.1 mm internal diameter) HPLC technology and microsequence analysis has enabled the positive identification of 125 of the total 179 amino acid residues (70%) in the molecule. The amino acid sequence data reported here permitted the isolation of a partial cDNA clone encoding LIF [Gearing et al. (1987) EMBO J. 6, 3995-4002]. A candidate C-terminus of the LIF molecule predicted from the amino acid sequence was confirmed by subsequent isolation of a cDNA clone corresponding to the C-terminus of the protein. No strong similarity was revealed when the amino acid sequence of LIF was compared with other haemopoietic growth factors, in particular granulocyte-macrophage colony-stimulating factor, granulocyte colony-stimulating factor and tumour necrosis factor-alpha or interleukins. The protein sequence data reported here indicate three sites of post-translational modification (N-linked glycosylation).