Biopolym. Cell. 2002; 18(5):436-441.
Molecular and Cell Biotechnologies
The influence of temperature on the yield soluble human alpha interferon in the system of recombinant proteins overproduction using bacteriophage lambda
1Slavchenko I. Yu.
  1. PSRS "Biotechnolog"
    150, Akademika Zabolotnogo Str., Kyiv, Ukraine, 03680


The recombinant proteins synthesized in E. coli cells are in need of their extraction from the host cells. Using bacteriophage lambda for infection of E. coli cells harbouring plasmid with target gene is one of the way of solving of this problem. As a result of lysis of E. coli cells by phage intracetlular target protein release to the growth medium. However, phage is able to enter either of two alternative modes of development upon infection of E. coli – lytic and lysogenic pathways. The different conditions affect on choose the lytic pathway. The influence of cultivation temperature (37, 28 and 21 OC) of E. coli cells harbouring plasmid with human alpha interferon gene infected by phage A on yield soluble interferon (IFN) in growth medium have been investigated. In this study it was shown that efficiency of bacteriophage lambda using for lysis producer and release to the growth medium soluble IFN depends on temperature of cultivation of infected cells. The maximum concentration of soluble target protein in cultural medium was observed at cultivation temperature 21 OC which is not optimal for lytic pathway of phage lambda development. The possible reasons of this result have been discussed.


[1] Pines O, Inouye M. Expression and secretion of proteins in E. coli. Mol Biotechnol. 1999;12(1):25-34.
[2] Blight MA, Holland IB. Heterologous protein secretion and the versatile Escherichia coli haemolysin translocator. Trends Biotechnol. 1994;12(11):450-5.
[3] Sandkvist M, Bagdasarian M. Secretion of recombinant proteins by Gram-negative bacteria. Curr Opin Biotechnol. 1996;7(5):505-11.
[4] Debabov DB. [Heterologous secretion in the Escherichia coli system]. Mol Biol (Mosk). 1994;28(3):496-505.
[5] Moir A, Brammar WJ. The use of specialised transducing phages in the amplification of enzyme production. Mol Gen Genet. 1976;149(1):87-99.
[6] Drew RE, Clarke PH, Brammar WJ. The construction in vitro of derivatives of bacteriophage lambda carrying the amidase genes of Pseudomonas aeruginosa. Mol Gen Genet. 1980;177(2):311-20.
[7] AS SSSR N 600175. Method of biosynthesis of biologically active substances. VA Kordium, SI Chernykh. Publ. in BI. N 12, 1978.
[8] Slavchenko IYu, Chernykh SI, Kordium VA. The study phagedependent synthesis b-lactamase in isogenic and Su Su + E. coli strains during infection of phages Xbla and XblaQ'R'. Enzymes of microorganisms. M.: VNII-SENTI, 1989; 31-6.
[9] Steffen D, Schleif R. Overproducing araC protein with lambda-arabinose transducing phage. Mol Gen Genet. 1977;157(3):333-9.
[10] Weiss B, Jacquemin-Sablon A, Live TR, Fareed GC, Richardson CC. Enzymatic breakage and joining of deoxyribonucleic acid. VI. Further purification and properties of polynucleotide ligase from Escherichia coli infected with bacteriophage T4. J Biol Chem. 1968;243(17):4543-55.
[11] Shub DA, Casna NJ. Bacteriophage T4, a new vector for the expression of cloned genes. Gene. 1985;37(1-3):31-6.
[12] Morita M, Asami K, Tanji Y, Unno H. Programmed Escherichia coli cell lysis by expression of cloned T4 phage lysis genes. Biotechnol Prog. 2001;17(3):573-6.
[13] Bl?si U, Kalousek S, Lubitz W. A bifunctional vector system for controlled expression and subsequent release of the cloned gene product by phi X174 lysis protein-E. Appl Microbiol Biotechnol. 1990;33(5):564-8. PubMed PMID: 6457237.
[14] Garrett J, Fusselman R, Hise J, Chiou L, Smith-Grillo D, Schulz J, Young R. Cell lysis by induction of cloned lambda lysis genes. Mol Gen Genet. 1981;182(2):326-31.
[15] Kloos DU, Str?tz M, G?ttler A, Steffan RJ, Timmis KN. Inducible cell lysis system for the study of natural transformation and environmental fate of DNA released by cell death. J Bacteriol. 1994;176(23):7352-61.
[16] Kalousek S, Schrot G, Lubitz W, Bl?si U. Expression of the Alcaligenes eutrophus phbA gene in Escherichia coli using a positive selection vector based on phage Lambda lysis genes. J Biotechnol. 1994;33(1):15-9.
[17] Srivastava R, Ali SS, Srivastava BS. Cloning of xylanase gene of Streptomyces flavogriseus in Escherichia coli and bacteriophage lambda-induced lysis for the release of cloned enzyme. FEMS Microbiol Lett. 1991;62(2-3):201-5.
[18] Pat. GB 2143238A. A method for enzyme liberation from bacterial cells. A. S. Breeze. Publ. date 06.02.1985.
[19] Pat USA 4,637,980. Externalization of product of bacteria. J. I. Auerbach, M. Rosenberg. Publ. date 20.01.1987.
[20] Slavchenko IYu. Study bacteriophage L efficiency for human a2 interferon production in Escherichia coli. Auth. Thesis. ... kand biol nauk. Kiev, 1990. 19 p.
[21] Slavchenko IYu. Isolation of clones sensitive to bacteriophage ? from the phage resistant Escherichia coli strain. Biopolym Cell. 2001; 17(2):160-5.
[22] Slavchenko IYu. The expression of human alpha-2b interferon in different strains of Escherichia coli. Biopolym Cell. 2001; 17(6):546-50.
[23] Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970;227(5259):680-5.
[24] AS. SSSR N 1092176. A method for preparing an artificial human interferon gene al and method for producing a polypeptide with an interferon activity microbiological synthesis. MN Kolosov, VG Korobko, VN Dobrynin, IV Severtsova, SA Chuvpilo, NS Bystrov, YuA Berlin, AL Kayushin, VV Butkus, IA Polyakova, EF Boldyreva, LS Sandakchiev, SG Popov, TN Shubina, VV Kravchenko, OI Serpinskiy, VF Yamschikov, SI Belikov, AN Sinyakov, GF Sivolobova. Publ. in BI N 18, 1984.
[25] Ptashne M. A Genetic Switch: Gene Control and Phage A. Cell Press, Cambridge, MA, and Blackwell Scientific, Palo Alto, CA, 1986. 138 pp.
[26] Schein CH. Production of Soluble Recombinant Proteins in Bacteria. Nat Biotechnol. 1989;7(11):1141–9.
[27] Obuchowski M, Shotland Y, Koby S, Giladi H, Gabig M, Wegrzyn G, Oppenheim AB. Stability of CII is a key element in the cold stress response of bacteriophage lambda infection. J Bacteriol. 1997;179(19):5987-91.
[28] Giladi H, Goldenberg D, Koby S, Oppenheim AB. Enhanced activity of the bacteriophage lambda PL promoter at low temperature. FEMS Microbiol Rev. 1995;17(1-2):135-40.
[29] Walker JR, Ussery CL, Allen JS. Bacterial cell division regulation: lysogenization of conditional cell division lon - mutants of Escherichia coli by bacteriophage. J Bacteriol. 1973;113(3):1326-32.
[30] Truitt CL, Haldenwang WG, Walker JR. Interaction of host and viral regulatory mechanisms: effect of the ion cell division defect on regulation of repression by bacteriophage lambda. J Mol Biol. 1976;105(2):231-44.