Protein having cell growth stimulating action, composition thereof and method for producing the same
AssigneeSanwa Kagaku Kenyusho Co., Ltd.
InventorsSetsuro Fujii, Nobumoto Chikazawa, Teruo Arima, Masakazu Fukushima
Life of this patent
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Abstract
A protein effective in stimulating cell growth activity. The protein has a molecular weight of from 5,000 to about 160,000, is composed predominantly of neutral and acidic amino acids, contains a significant amount of glutamic acid and aspartic acid, and is substantially free of nucleoside phosphotransferase. The protein is useful as wound treatment agent and as promoting agent in the synthesis of DNA. A method of producing the protein and compositions containing the same are also disclosed.
Claims30
A protein having cell growth stimulating activity, said protein selected from those obtained from a culture of a microorganism selected from the group consisting of Staphylococcus epidermidis, Bacillus subtilis, Streptococcus faecalis, Serratia marcescens, Escherichia coli, Salmonella enteritidis, and Proteus vulgaris, the protein being characterized as follows:
- (a) Molecular weight in the range of 5,000-12,000, when said molecular weight is calculated on the basis of a calibration curve plotting the relationship between the molecular weight and an amount eluted by high performance liquid chromatography using as marker proteins, thioglobin (M.W. 670,000), bovine .gamma.-globulin (M.W. 158,000), fowl ovalbumin (M.W. 44,000), equine myoglobulin (M.W. 17,000), and vitamin B-12 (M.W. 1,350);
- (b) Ultraviolet absorption at 278 nm;
- (c) Infrared absorption spectrum with dominant peaks at 1300, 1550, and 1630 cm.sup.-1 ;
- (d) Isoelectric point in the range of 4.0-6.6;
- (e) Soluble in water and insoluble in methanol, ethanol, acetone and ether;
- (f) Appears as a grayish white powder in the lyophilized state;
- (g) Positive for the following color reactions;
- Xanthoprotein, Pauli, Sakaguchi, Hunter, Sullivan, Hopkins-Cove, nitrosonaphthol, Millon, Neubauer-Road, and Libermann;
- and
- (h)
- ______________________________________ Amino Acid mol/M ______________________________________ Aspartic Acid 4.25-14.33 Glutamic Acid 4.76-21.35 Glycine 3.68-10.04 Valine 3.05-8.97 Alanine 3.51-18.09 Threonine 2.20-5.69 Serine 2.01-5.14 Cystine 0.00-0.40 Methionine 0.79-2.73 Isoleucine 2.34-7.97 Leucine 2.70-8.70 Tyrosine 0.39-3.17 Phenylalanine 0.79-3.20 Lysine 2.22-8.80 Histidine 0.54-1.35 Arginine 0.65-4.83 Proline 2.63-5.37 Tryptophan 0.25-0.70 ______________________________________
- said protein exhibiting cell growth stimulating activity in a pH range between 4-10;
- and being substantially free of nucleoside phosphotransferase.
A protein having cell growth stimulating activity, said protein selected from those obtained from a culture of a microorganism selected from the group consisting of Staphylococcus aureus, Sarcina lutea, Aerococcus viridians, Acinetobacter calcoaceticus, Corynebacterium fascians, and Clostridium perfringens;
- the protein being characterized as follows:
- (a) Molecular weight in the range 10,000-15,000, when said molecular weight is calculated on the basis of a calibration curve plotting the relationship between the molecular weight and an amount eluted by high performance liquid chromatography using as marker proteins, thioglobin (M.W. 670,000), bovine .gamma.-globulin (M.W. 158,000), fowl ovalbumin (M.W. 44,000), equine myoglobin (M.W. 17,000), and vitamin B-12 (M.W. 1,350);
- (b) Ultraviolet absorption at 278 nm;
- (c) Infrared absorption spectrum with dominant peaks at 1300, 1550, and 1630 cm.sup.-1 ;
- (d) Soluble in water and insoluble in methanol, ethanol, acetone and ether;
- (e) Appears as a grayish white powder in the lyophilized state;
- (f) Positive for the following color reactions:
- Xanthoprotein, Pauli, Sakaguchi, Hunter, Sullivan;
- Hopkins-Cole, nitrosonaphthol, Millon, Neubauer-Road, and Liebermann;
- (g) Isoelectric point in the range of 4.0-5.5;
- and
- (h)
- ______________________________________ Amino Acid mol/M ______________________________________ Aspartic Acid 9.65-18.78 Glutamic Acid 12.36-43.08 Glycine 6.71-11.78 Valine 6.43-12.16 Alanine 8.07-14.68 Threonine 4.60-7.72 Serine 4.56-11.45 Cystine 0.00 Methionine 1.46-2.86 Isoleucine 3.98-6.77 Leucine 4.97-8.13 Tyrosine 0.62-2.63 Phenylalanine 1.75-3.44 Lysine 4.53-8.95 Histidine 1.04-1.78 Arginine 0.80-2.33 Proline 4.27-12.69 Tryptophan 0.32-0.63 ______________________________________
- said protein being substantially free of nucleoside phosphotransferase.
A protein having cell growth stimulating activity, said protein selected from those obtained from a culture of the microorganism Clostridium perfringens, the protein being characterized as follows:
- (a) Molecular weight of 150,000-160,000, when said molecular weight is calculated on the basis of a calibration curve plotting the relationship between the molecular weight and an amount eluted by high performance liquid chromatography using as marker proteins, thioglobin (M.W. 670,000), bovine .gamma.-globulin (M.W. 158,000), fowl ovalbumin (M.W. 44,000), equine myoglobulin (M.W. 17,000), and vitamin B-12 (M.W. 1,350);
- and a molecular weight of approximately 53,000, when determined by SDS disk electrophoresis;
- (b) Ultraviolet absorption at 278 nm;
- (c) Infrared absorption spectrum with dominant peaks at 1300, 1550, and 1630 cm.sup.-1 ;
- (d) Soluble in water and insoluble in methanol, ethanol, acetone and ether;
- (e) Appears as a grayish white powder in the lyophilized state;
- (f) Positive for the following color reactions:
- Xanthoprotein, Pauli, Sakaguchi, Hunter, Sullivan;
- Hopkins-Cole, nitrosonaphthol, millon, Neubauer-Road, and Liebermann;
- and
- (g)
- ______________________________________ Amino Acid mol/M ______________________________________ Aspartic Acid 109.80 Glutamic Acid 96.18 Glycine 71.66 Valine 73.01 Alanine 73.69 Threonine 40.39 Serine 45.59 Cystine 2.92 Methionine 16.56 Isoleucine 54.64 Leucine 65.42 Tyrosine 54.29 Phenylalanine 27.43 Lysine 75.66 Histidine 10.48 Arginine 21.37 Proline 32.55 Tryptophan 3.36 ______________________________________
- said protein being substantially free of nucleoside phosphotransferase.
Dependencies are inferred from the claim text.