F. William Studier

Molecular & Structural Biology Group Leader

Research Interests

Research has centered on conformations and interactions of DNA, molecular genetics and biochemistry of bacteriophage T7, and making T7 RNA polymerase and T7 expression signals useful for production of RNAs and proteins

T7 has been a continuing interest since the 1960s. T7 genes were defined and mapped genetically, and functions of many of them have been determined by a variety of biochemical and physical techniques. Processes studied include entry of T7 DNA into its host cell, E. coli; overcoming host restriction; expression of T7 genes and shut-off of host functions; replication, processing and packaging of T7 DNA; and structure and assembly of phage particles. Determination of the complete nucleotide sequence of T7 DNA, 39,937 base pairs, revealed coding sequences for more than fifty T7 proteins and the arrangement of signals that direct gene expression. All of the T7 proteins can now be expressed from clones, independently or during infection, and virus mutants defective in any T7 gene have been isolated and characterized.

The challenges and outcomes of this basic research spurred the development of some widely used research methods, including sedimentation to measure size and shape of single- and double-stranded DNA, and slab gels for electrophoresis of proteins and nucleic acids. An understanding of how T7 efficiently directs an infected cell to produce T7 gene products led to the cloning of T7 RNA polymerase, a highly selective enzyme that can produce almost any RNA, and development of an inducible expression system that uses T7 RNA polymerase and T7 expression signals in pET vectors to produce proteins from cloned coding sequences in E. coli BL21(DE3).

Subsequent work has concentrated on improving the convenience and reliability of the T7 expression system. Non-inducing growth media have been developed for stable maintenance of expression strains, and auto-inducing media have been developed that allow production of target proteins simply by inoculating with the expression strain and growing to saturation. These media and methods increase the convenience and reliability of protein production at any scale but are particularly useful for high throughput, because they facilitate protein production in many cultures in parallel and generally produce more target protein per volume of culture than is obtained in conventional media by induction with IPTG. PDFs may be downloaded here of an Overview & Recipes for non-inducing and auto-inducing media and a chapter “Stable Expression Clones and Auto-Induction for Protein Production in E. coli” submitted for the volume Structural Genomics: Methods and Protocols, edited by Yu Wai Chen for the series Methods in Molecular Biology."

Recent collaborations have traced the origin of E. coli B, a progenitor of BL21(DE3), and determined the genome sequences of BL21(DE3) and REL606, another B strain used by Richard Lenski for evolutionary studies. Every difference between these two genome sequences can be given a reasonable explanation in light of their published and inferred laboratory histories. Detailed comparisons were also made between the genome sequences of B and K-12, two isolates of commensal E. coli that came to prominence in the 1940s for studies of bacteriophages and biochemical genetics. Derivatives of E. coli B and K-12 are the most widely studied strains of laboratory bacteria and work with them has provided much of our understanding of molecular genetics. A current interest is to compare the genome sequences of B and K-12 with those of the burgeoning number of isolates of both commensal and pathogenic E. coli whose genomes have been sequenced, to gain insights into bacterial evolution.

Recent News

• F. William Studier: Basic Research Leads to Most Successful BNL Technology
• Genomes of Two Popular Research Strains of E. coli Sequenced

Selected Publications

• Daegelen P., Studier F.W., Lenski R. E., Cure S., and Kim J.F.
  Tracing ancestors and relatives of Escherichia coli B, and the derivation of B strains REL606 and BL21(DE3).
  Journal of Molecular Biology, 394(4):634-643 (2009).  PubMed
• Jeong H., Barbe,V., Lee C. H., Vallenet D., Yu D. S., Choi S., Couloux A., Lee S., Yoon S. H., Cattolico L., Hur C., Park H., Segurens B., Kim S.C., Oh T. K., Lenski R. E., Studier F.W., Daegelen P., and Kim J. F.
  Genome sequences of Escherichia coli B strains REL606 and BL21(DE3).
  Journal of Molecular Biology, 394(4):644-652 (2009).  PubMed
• Studier F.W., Daegelen P., Lenski R.E., Maslov S., and Kim J.F.
  Understanding the differences between genome sequences of Escherichia coli B strains REL606 and BL21(DE3) and comparison of the E. coli B and K-12 genomes.
  Journal of Molecular Biology, 394(4):653-680 (2009).  PubMed
• Bewley M.C., Graziano V., Jiang J., Matz E., Studier F.W., Pegg A.E., Coleman C.S. and Flanagan J.M.
  Structures of wild-type and mutant human spermidine/spermine N1-acetyltransferase, a potential therapeutic drug target.
  Proc Natl Acad Sci USA, 103:2063-2068 (2006).  PubMed  Full Text
  PDB Files:2B5G   2B4D   2B3U   2B3V   2B58   2B4B  
• Studier F.W.
  Protein production by auto-induction in high-density shaking cultures.
  Protein Expr Purif., 41:207-234 (2005).  PubMed
• Zhang X. and Studier F.W.
  Multiple roles of T7 RNA polymerase and T7 lysozyme during bacteriophage T7 infection.
  J Mol Biol., 340:707-730 (2004).  PubMed

• Studier F.W.
  Slab gel electrophoresis.
  Trends Biochem Sci., 25:588-590 (2000).  PubMed  Full Text
• Burley S.K., Almo S.C., Bonanno J.B., Capel M., Chance M.R., Gaasterland T., Lin D., Sali A., Studier F.W. and Swaminathan S.
  Structural genomics: beyond the human genome project.
  Nature Genetics, 23:151-157 (1999).  PubMed  Full Text
  
• Zhang X. and Studier F.W.
  Mechanism of inhibition of bacteriophage T7 RNA polymerase by T7 lysozyme.
  J Mol Biol., 269:10-27 (1997).  PubMed

• Cerritelli M.E. and Studier F.W.
  Assembly of T7 capsids from independently expressed and purified head protein and scaffolding protein.
  J Mol Biol., 258:286-298 (1996).  PubMed
• Goldman E., Rosenberg A.H., Zubay G. and Studier F.W.
  Consecutive low-usage leucine codons block translation only when near the 5' end of a message in Escherichia coli.
  J Mol Biol., 245:467-473 (1995).  PubMed
• Cheng X., Zhang X., Pflugrath J.W. and Studier F.W.
  The structure of bacteriophage T7 lysozyme,
  a zinc amidase and an inhibitor of T7 RNA polymerase.
  Proc Natl Acad Sci USA, 91:4034-4038 (1994).  PubMed  Full Text
  PDB File:1LBA  
• Rosenberg A.H., Goldman E., Dunn J.J., Studier F.W. and Zubay G.
  Effects of consecutive AGG codons on translation in Escherichia coli, demonstrated with a versatile codon test system.
  J. Bacteriol., 175:716-722 (1993)  PubMed
• Studier F.W., Rosenberg A.H., Dunn J.J. and Dubendorff J.W.
  Use of T7 RNA polymerase to direct expression of cloned genes.
  Methods in Enzymology, 185:60-89 (1990).  PubMed
• Studier F.W., and Bandyopadhyay P.K.
  Model for how type I restriction enzymes select cleavage sites in DNA.
  Proc Natl Acad Sci USA, 85:4677-4681 (1988).  PubMed  Full Text
  
• Moffatt B.A. and Studier F.W.
  Entry of bacteriophage T7 DNA into the cell and escape from host restriction.
  J Bacteriol, 170:2095-2105 (1988).  PubMed
• Moffatt B.A. and Studier F.W.
  T7 lysozyme inhibits transcription by T7 RNA polymerase.
  Cell, 49:221-227 (1987).  PubMed
• DeMassy B.,Weisberg R.A. and Studier F.W.
  Gene 3 endonuclease of bacteriophage T7 resolves conformationally branched structures in double-stranded DNA.
  J Mol Biol, 193:359-376 (1987).  PubMed
• Studier F.W. and Moffatt B.A.
  Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes.
  J Mol Biol, 189:113-130 (1986).  PubMed

• Davanloo P., Rosenberg A.H., Dunn J.J. and Studier F.W.
  Cloning and expression of the gene for bacteriophage T7 RNA polymerase.
  Proc Natl Acad Sci USA, 81:2035-2039 (1984).  PubMed  Full Text
  
• Studier F.W. and Dunn J.J.
  Organization and expression of bacteriophage T7 DNA.
  Cold Spring Harbor Symp Quant Biol., 47:999-1007 (1983).  PubMed
• Dunn J.J. and Studier F.W.
  Complete nucleotide sequence of bacteriophage T7 DNA and the locations of T7 genetic elements.
  J Mol Biol, 166:477-535 (1983).  PubMed    NCBI Sequence File of T7 DNA
• Studier F.W.
  Relationships among different strains of T7 and among T7-related bacteriophages.
  Virology, 95(1):70-84 (1979).  PubMed
• McDonell M.W., Simon M.N. and Studier F.W.
  Analysis of restriction fragments of T7 DNA and determination of molecular weights by electrophoresis in neutral and alkaline gels.
  J Mol Biol., 110:119-146 (1977).  PubMed

• Studier F.W.
  Gene 0.3 of bacteriophage T7 acts to overcome the DNA restriction system of the host.
  J Mol Biol., 94:283-295 (1975).  PubMed
• Dunn J.J. and Studier F.W.
  T7 early RNAs are generated by site-specific cleavages.
  Proc Natl Acad Sci USA, 70:1559-1563 (1973).  PubMed  Full Text
  

• Studier F.W.
  Analysis of bacteriophage T7 early RNAs and proteins on slab gels.
  J Mol Biol, 79:237-248 (1973).  PubMed
• Studier F.W.
  Bacteriophage T7.
  Science, 176:367-376 (1972).  PubMed

• Studier F.W.
  Effects of the conformation of single-stranded DNA on renaturation and aggregation.
  J Mol Biol, 41:199-209 (1969).  PubMed
• Studier F.W.
  Sedimentation studies of the size and shape of DNA.
  J Mol Biol, 11:373-390 (1965).