Amino acid sequences of alpha-helical segments from S-carbosymethylkerateine-A. Complete sequence of a type-I segment.

The amino acid sequence of a type-I helical segment from the low-sulphur protein (S-carboxymethylkerateine-A) of wool was determined by combining automatic and manual-sequencing data. Whereas in the type-II helical segment most of the cationic groups occur in pairs, 11 of the 22 anionic residues in the sequence of the type-I segment were situated next to a second anionic residue. This suggests possible interactions between type-I and type-II helical segments in alpha-keratin. As observed with the sequence of a type-II helical segment a model constructed on 3.6 residues per turn of helix shows a line of hydrophobic residues along the helix, thereby supporting the physicochemical evidence that the molecule is predominantly helical and forms part of a coiled-coil structure. Examination of the sequence data by predictive methods indicates the possibilty of extensive sections of alpha-helix interspersed with discontinuities. The molecule contains a number of regions with peptide sequences identical with those found by other workers after enzymic digestion of fractions from oxidized wool.     

Complete nucleotide sequence of a porcine HSP70 gene

The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession number M69100.     

Aggregation of wool keratin intermediate filament proteins

The wool keratin intermediate filament proteins were isolated as their S-carboxymethyl derivatives (S-carboxymethylkerateine A, SCMKA) and purified by gel filtration to remove residual non-helical protein of low molecular weight. The alpha-helix content of purified SCMKA was approximately 62% in agreement with that predicted for the alpha-helical coiled-coil segments from the amino acid sequences of the subunits. In aqueous buffer at pH 11 or in n-propanol (20% v/v) at pH 9.2 very large aggregates are dissociated and SCMKA exists largely as a mixture of the dimer (two-chain coiled-coil of Mr approximately 103,000) and the tetramer. The protein species are not in rapidly reversible equilibrium as judged from gel filtration and sedimentation equilibrium. It is probable that species with a range of association constants are present. The equilibrium is shifted towards the dimer with change of pH from 9.2 to 11 or by the addition of 20% (v/v) n-propanol. The tetrameric proteolytic digestion product which is derived from the 1B segment of the alpha-helical rod section of the keratin molecule dissociates in a similar way to intact SCMKA with increase of pH and in the presence of n-propanol. This indicates the importance of this region of the rod domain in the initial stages of the assembly of the filament. Electrostatic and hydrophobic interactions are implicated in the association of the two-chain coiled-coil to the tetramer both in intact SCMKA and the 1B segment tetramer. The results are discussed in relation to the intact dimeric and tetrameric complexes obtained from other intermediate filament types.     

The amino acid sequence of protein SCMK-B2A from the high-sulphur fraction of wool keratin

1. The amino acid sequence of protein SCMK-B2A, a reduced and S-carboxymethylated protein from the high-sulphur fraction of wool, has been determined. 2. This protein of 171 amino acid residues displays both a high degree of internal homology and extensive external homology with other members of the SCMK-B2 group of proteins. 3. Evidence is presented which suggests that the SCMK-B2 group of proteins are produced by separate non-allelic genes.     

Evidence of linkage between high-glycine-tyrosine keratin gene loci and wool fibre diameter in a Merino half-sib family

Candidate genes for quantitative trait loci have been studied in a Medium Peppin Merino flock. Obvious candidates for effects on wool production traits are genes for the major proteins expressed in the wool fibre, the keratin and keratin-associated protein genes. Two keratin-associated protein loci, KRTAP6 and KRTAP8, have previously been shown to be linked. The results of analyses between these two loci and production traits gave significant evidence of linkage with wool fibre diameter in one out of eight halfsib groups tested. High-glycine-tyrosine proteins (KRTAP6, 7 and 8) are known to vary considerably in abundance in wool fibres and it is possible that a gene for major effect on fibre diameter is located within the same chromosomal region as KRTAP6 and KRTAP8.     

The amino acid sequence of protein SCMK-B2C from the high-sulphur fraction of wool keratin

1. The amino acid sequence of a protein from the reduced and carboxymethylated high-sulphur fraction of wool has been determined. 2. The sequence of this S-carboxymethylkerateine (SCMK-B2C) of 151 amino acid residues displays much internal homology and an unusual residue distribution. Thus a ten-residue sequence occurs four times near the N-terminus and five times near the C-terminus with few changes. These regions contain much of the molecule's half-cystine, whereas between them there is a region of 19 residues that are mainly small and devoid of cystine and proline. 3. Certain models of the wool fibre based on its mechanical and physical properties propose a matrix of small compact globular units linked together to form beaded chains. The unusual distribution of the component residues of protein SCMK-B2C suggests structures in the wool-fibre matrix compatible with certain features of the proposed models.     

Sheep wool (glycine + tyrosine)-rich keratin genes. A family of low sequence homology

We report the isolation and complete nucleotide sequence of genes encoding the two major high-(glycine + tyrosine) (HGT) keratins of sheep wool in separate genomic clones. The genes have negligible sequence homology except for an 18-bp conserved element immediately preceding the initiation codon. The same element has been found in the corresponding position of a number of co-expressed high-sulphur (HS) keratin genes and may therefore represent a common element between these wool intermediate-filament-associated proteins. As seen for the HS keratin genes, the HGT keratin genes also lack introns. Southern blot data show that both HGT genes are unique in the sheep genome, indicating that the observed heterogeneity of the HGT protein class may not be as complex as previously suggested.     

Complete sequence and gene organization of the Nosema spodopterae rRNA gene

By sequencing the entire ribosomal RNA (rRNA) gene of Nosema spodopterae, we show here that its gene organization follows a pattern similar to the Nosema type species, Nosema bombycis, i.e. 5'-large subunit rRNA (2,497 bp)-internal transcribed spacer (185 bp)-small subunit rRNA (1,232 bp)-intergenic spacer (277 bp)-5S rRNA (114 bp)-3'. Gene sequences and the secondary structures of large subunit rRNA, small subunit rRNA, and 5S rRNA are compared with the known corresponding sequences and structures of closely related microsporidia. The results suggest that the Nosema genus may be heterogeneous and that the rRNA gene organization may be a useful characteristic for determining which species are closely related to the type species.