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R C Lu

Publications and source records attributed to R C Lu.

28 records · Page 2Linked to original sources
PubMed1983

The proteolytic substructure of light meromyosin. Localization of a region responsible for the low ionic strength insolubility of myosin.

Light meromyosin (LMM), prepared by limited tryptic digestion of myosin, usually contains several polypeptide chains, LMM-A, LMM-B, and LMM-C in decreasing order of molecular weight estimated from sodium dodecyl sulfate-gel electrophoresis. Further limited tryptic digestion of LMM produces well defined fragments (Balint, M., Szilagyi, L., Fekete, Gy., Blazso, M., and Biro, E. N. A. J. Mol. Biol. (1968) 37, 317-330). Fragments LF-1, LMM-D, LF-2, and LF-3, with chain masses equal to 63, 56, 47, and 30 kDa, respectively, have been isolated by column chromatography. Based on the time course of the changes in the sodium dodecyl sulfate-gel pattern of the digests, chain masses estimated from sodium dodecyl sulfate-gel electrophoresis, and the NH2- and COOH-terminal sequences of the isolated peptides, the following scheme can be deduced. Formula; see text. C and N over the arrows indicate removal of residues from the COOH and NH2 terminus, respectively. The positions of the peptides along the myosin heavy chain have been established by comparison with the published primary structures of rabbit skeletal (Elzinga, M., Behar, K., Walton, G., and Trus, B. L. (1980) Fed. Proc. 33, 1579) and nematode myosin (McLachlan, A. D., and Karn, J. (1982) Nature (Lond.) 299, 226-231). LMM and fragment LMM-D are insoluble, whereas LF-1, LF-2, and LF-3 are soluble at low ionic strength. Their solubility properties, in conjunction with their locations along the myosin heavy chain, suggest that a relatively small stretch of peptide (chain weight, 5,000 Da) located about 100 residues from the COOH terminus of myosin heavy chain is responsible for the insolubility of LMM at low ionic strength.

Animals↗
PubMed1982

Changes of lysine reactivities of actin in complex with myosin subfragment-1, tropomyosin and troponin.

The interactions of actin with myosin subfragment-1 and tropomyosin were explored by comparing the reactivities of lysine residues in F-actin alone with those in F-actin complexed to the other proteins. Limited reductive methylation was carried out on F-actin and the F-actin complexes with [14C]HCHO and [3H]HCHO, respectively. After dissociation from the other components, [3H]actin was combined with [14C]actin and 3H/14C of each lysine residue was measured. Myosin subfragment-1 reduced the reactivities of Lys-335 and Lys-372, while tropomyosin reduced those of Lys-237, -325, 327 and -335. When troponin was present in the absence of Ca2+, the effect of tropomyosin on Lys-335 remained the same, but its reactivity was completely restored upon the addition of Ca2+. Thus, the results suggest that different parts of actin are affected by the interaction with myosin subfragment-1 and tropomyosin, but the region containing Lys-335 is commonly affected by the presence of either of them. The change in reactivity is attributable either to a direct steric effect or to an induced conformational effect.

Actins↗
PubMed1981

Digestion of troponin C with trypsin in the presence and absence of Ca2+. Identification of cleavage points.

The rate of tryptic digestion of troponin C has been shown to be dependent on Ca2+ (Drabikowski et al., Biochim. Biophys. Acta 490, 216-224). We have characterized the tryptic peptides produced both in the presence and absence of Ca2+ using amino acid composition and end-group analyses. In the presence of Ca2+ trypsin cleaves TnC at Arg-8, Lys-84 and Lys-88, leading to the formation of two large peptides, one containing the two low-affinity sites (TR1C), the other, the two high-affinity Ca2+-binding sites (TR2C). In the absence of Ca2+ (1 mM EDTA), digestion proceeds much more rapidly and takes place first at Arg-100, followed by Arg-104, Arg-120, Lys-153, Arg-8 and others. The data suggest that the points of cleavage are determined by the Ca2+-dependent conformational states of TnC, particularly in the C-terminal half of the protein where the cation is known to induce secondary structure.

Amino Acid Sequence↗
PubMed1981

Change of reactivity of lysine residues upon actin polymerization.

The reactivity of lysine residues of actin was measured by a surface labeling method--limited reductive methylation. After labeling, actin was subjected to CNBr and enzymatic cleavage, and all lysines were obtained either singly in a peptide or as a free residue. The specific activity of each lysine was taken as the measure of its reactivity. In actin denatured in 8 M urea, the reactivity of each lysine residue is approximately equal whereas those in G-actin fall into three categories: Lys-61 and Lys-113 are the most reactive ones; Lys-18, -213, -215, -314, and -358 are hardly reactive; the remainder, including Lys-50, -68, -84, -118, -191, -237, -283, -290, -325, -327, -335, and -372, are moderately reactive. The least reactive ones are probably buried in the native G-actin and all the others are most likely on the surface. Upon actin polymerization the reactivities of Lys-61, -68, -113, and -283 are significantly reduced while that of Lys-335 is strikingly enhanced. The decrease in reactivity could be readily explained if these residues were located in the monomer-monomer contact area although a polymerization-induced conformational change cannot be excluded. Such a conformational change may be invoked to explain the increase in the reactivity of Lys-335. Alternatively, the latter may be interacting with the bound ATP of G-actin, and the increased reactivity might be directly attributable to the loss of gamma-P for ATP accompanying polymerization.

Actins↗
PubMed1980

Identification of a region susceptible to proteolysis in myosin subfragment-2.

Comparison of the NH2-terminal sequence of myosin short subfragment-2 (Mr of subunit = 37,000) and long subfragment-2 (Mr of subunit = 59,000) demonstrates that the former represents the NH2-terminal portion of the latter and suggests that the hinge region in myosin rod is in the COOH-terminal portion of the long subfragment-2.

Amino Acid Sequence↗
PubMed1978

The primary structure of tubulin. Sequences of the carboxyl terminus and seven other cyanogen bromide peptides from the alpha-chain.

The alpha-chain of calf brain tubulin was fragmented by treatment with cyanogen bromide and eight peptides together accounting for 108 residues were purified and sequenced. The COOH-terminal peptide contains a fractional amount (about 0.3 residues) of tyrosine at its COOH-terminus; this presumably represents tyrosine that is added post-translationally to alpha-tubulin. The beta-chain can be phosphorylated, and the probable site of this modification is identified also in the COOH-terminal peptide. Comparison of the sequences described here with the sequence of actin reveals no homology between actin and tubulin.

Amino Acid Sequence↗
PubMed1978

Cleavage of a specific bond in troponin C by thrombin.

Limited proteolysis of rabbit skeletal troponin C with bovine thrombin yielded two fragments, TH1 (Mr = 11000) containing Ca2+ binding regions I--III and TH2 (Mr = 6000) containing region IV. Determination of the partial sequences of the fragments established the site of cleavage at Arg120-Ala121. Secondary cleavage by thrombin at other arginyl or lysyl residues in troponin C was ruled out by the sequence data and by the amino acid compositions of the two fragments.

Alanine↗
PubMed1977

Partial amino acid sequence of brain actin and its homology with muscle actin.

Actin was purified from calf brains by chromatography on DEAE-Sephadex and hydroxylapatite. The protein was then subjected to amino acid sequence analysis by isolating and sequencing its cyanogen bromide peptides. CB-1, 3, 4, 5, 6, 9, 10, and 12 correspond to equivalent segments of rabbit skeletal muscle actin, while subsitutions involving methionines give rise to some new peptides. The region that corresponds to CB-13 in muscle actin becomes two peptides in the brain protein because of a Leu leads to Met replacement at position 16, while Met leads to Leu substitutions at positions 176 and 298 give rise to two larger peptides, CB-15 + 7 and CB-8 + 2, which correspond to muscle actin CB-15 fused with CB-7 and CB-8 fused with CB-2, respectively. The peptides that have been isolated from brain actin contain 267 of the 374 residues in actin, of which 157 have been unequivocally identified. When the data are compared with those for rabbit skeletal muscle actin, 11 replacements are seen; thus the two actins differ at about 7% of the positions examined.

Actins↗