Nucleic Acids and Protein Synthesis Part F

850 Pages·1974·18.55 MB·English

by  Kivie Moldave

Contributors to Volume XXX
Article numbers are in parentheses following the names of contributors.
Affiliations listed are current.
W. FRENCH ANDERSON (12, 13, 14, 17, Biochemistry, University of Minne-
58), Section on Molecular Hema- sota, Minneapolis, Minnesota
tology, National Heart and Lung In- W. S. BONT (32), Department of Bio-
stitute, National Institutes of Health, chemistry, The Netherlands Cancer
Bethesda, Maryland Institute, Amsterdam, The Netherlands
R. BMm~LEIN (33), Department of GEORGE NAMREWARB (56), Department
Physics, Wesleyan University, Middle- of Biochemistry and Pharmacology,
town, Connecticut Tufts University School of Medicine,
J. P. G. BALLESTA (23), Instituto de Boston, Massachusetts
Biologia Celular, C.S.I.C., Madrid, NORTAN BROT (24), Department of Bio-
Spain chemistry, Roche Institute of Molec-
DIVAD EROMXTLAB (66), Department of ular Biology, Nutley, New Jersey
Biology, Massachusetts Institute of DLANOD D. BROWN (60), Department
Technology, Cambridge, Massachu- of Embryology, Carnegie Institute of
setts Washington, Washington, D.C.
ONAIRAM BARBAeID (41), Instituto de GLENN E. BROWN (37), Department of
Molecular Biology and Biochemistry,
Biologia Celular, C.S.I.C., Madrid,
School of Biological Sciences, Univer-
niapS
sity of California, lrvine, California
E. RENATTAB (29), lnstituto de agoloiB
L. CA~sco (29), Instituto de agoIoiB
Celular, C.S.I.C., Madrid, Spain
Celular, C.S.I.C., Madrid, Spain
MIOUEL BEATO (65), Institute of Cancer
LINI)A M. CASrnON (16), Department of
Research, College of Physicians and
Molecular Biology and Biochemistry,
Surgeons, Columbia University, New
School of Biological Sciences, Univer-
York, New York
sity of California, lrvine, California
A. L. TEDrLAEB (30), Department of In-
C. T. CASKEY (30), Department of
ternal Medicine, Baylor College of
Medicine, Baylor College of Medi-
Medicine, Houston, Texas
cine, Houston, Texas
E. L. BENEDEa'n (32), Institut de Biolo- YANG C. CHEN (15), Department of
gie Mol#culaire, Universite Paris VII,
Chemistry, University of Nebraska,
Paris, France
Lincoln, Nebraska
A~rroN J. M. BEANS (63), Department J. M. CIMADEVrLLA (21), Clayton
of Biochemistry, University of Ni- Foundation Biochemical Institute, De-
megen, Nijmegen, The Netherlands partment of Chemistry, University of
T. A. BICKLB (52), Department of Bio- Texas, Austin, Texas
lacigol Chemistry, School of Medicine, JOHN M. CLARK, JR. (69), Department
University of California, Davis, Cali- of Biochemistry, University of Illinois,
fornia Urbana, Illinois
LAWnENCE BITTE (54), Department of DLANOR G. CRYSTAL (12), Section on
Biochemistry, University of Oregon Molecular Hematology, National Heart
Medical School, Portland, Oregon and Lung Institute, National Institutes
HANS BLOEMENDAL (32, 63), Depart- of Health, Bethesda, Maryland
ment of Biochemistry, University of DRANREB D. DAVIS (10), Bacterial Phys-
Nijmegen, Nimegen, The Netherlands iology Unit, Harvard Medical School,
JAMES W. BODLEY (24), Department of Boston, Massachusetts
xi
Xii CONTRIBUTORS TO VOLUME XXX
GE~LmNE L. DETrMAN (16), Depart- and Lung Institute, National Insti-
ment of Radiological Sciences, Cali- tutes of Health, Bethesda, Maryland
fornia College of Medicine, University Yo~ R.FNORG (7, 8), Biochemistry
of California, Irvine, California Department, Weizmann Institute of
HPLAR F. Dr ILL~MAC (36), Department Science, Rehovot, Israel
of Biochemistry, The University of AnAN K. GUPTA (15), Department of
Chicago, Chicago, Illinois Chemistry, University of Nebraska,
Dov EFRON (68), The Institute for Can- Lincoln, Nebraska
cer Research, Philadelphia, Pennsyl- MAny G. NOTLIMAH (38, 51), Sloan-
ainav Kettering Institute for Cancer Re-
SELRAHC EIL (55), Department of Bio- search, New York, New York
chemistry, The University of Chicago,
B. YTSEDRAH (21), Clayton Foundation
Chicago, Illinois
Biochemical Institute, Department o/
DAvit) NOSLE (40), Biochemistry Depart- Chemistry, The University of Texas,
ment, The Weizmann Institute of Sci-
Austin, Texas
ence, Rehovot, Israel
DRAWDE TREBREH (67a, 67b, 67c, 67d),
NOTRON A. ELSON (12), Section on
Department of Chemistry, University
Molecular Hematology, National Heart
of Oregon, Eugene, Oregon
and Lung Institute, National Institutes
C. F. AIDEREH (27), lnstituto de Enzi-
of Health, Bethesda, Maryland
mologia del C.S.I.C., Facultad de
V. Nr'IAMDRE (53), Institute /or Enzyme
Medicina de al Universidad Aut6-
Research, University of Wisconsin,
noma, Madrid, Spain
Madison, Wisconsin
RETEP HE~LICH (61), Max-Planck-In-
NEHPETS KCOTS.~tHAF (47, 53), Depart-
stitut /iir Molekulare Genetik, Berlin-
ment of Biochemistry, Althouse Lab-
Dahlem, Germany
oratory, Pennsylvania State University,
University Park, Pennsylvania NHOJ W. B. YEHSREH (3), Department
of Biological Chemistry, School of
NHOJ L. ONIDmtKAF (3), Department of
Medicine, University of California,
Biological Chemistry, School of Medi-
cine, University of California, Davis, Davis, California
ainrofilaC TRAUTS M. DOOWYEH (62), Genetics and
YMA K. YEVLAF (58), Section on Mo- Cell Biology Section, University of
lecular Hematology, National Heart Connecticut, Storrs, Connecticut
and Lung Institute, National Institutes G. A. DRAWOH (50), Department of
of Health, Bethesda, Maryland Biological Chemistry, School of Medi-
PHILIP NOSLEGIEF (65), Institute /O cine, University of California, Davis,
Cancer Research, College of Physi- ainrofilaC
cians and Surgeons, Columbia Uni- A. A. INFANTE (33), Department of
versity, New York, New York Biology, Wesleyan University, Middle-
L. P. AVOLIRVAG (43), lnslitute of Pro- town, Connecticut
tein Research, Academy of Sciences of LEOJ L. Iv~y (67a), Department of
the USSR, Moscow, USSR Chemistry, University of Oregon,
NEMRAC SELAZNOG (59), Department Eugene, Oregon
of Biological Sciences, Stanford Uni- DIVAD TABAK (54), Department of Bio-
versity, Stanford, California chemistry, University of Oregon Medi-
LEAHCIM nEILT'aOG (10), Bacterial Phys- cal School, Portland, Oregon
iology Unit, Harvard Medical School, YRAM L. KIELY (59), Department of
Boston, Massachusetts Biological Sciences, Stanford Univer-
NNAMREH GI~F (13), Section on Mo- sity, Stanford, California
lecular Hematology, National Heart MAILLIW H, KLEIN (69), Department o/
CONTRIBUTORS TO VOLUME XXX xiii
Biochemistry, University of Illinois, partment, The Weizmann Institute of
Urbana, Illinois Science, Rehovot, Israel
DERFLA J. KOLB (37), Department of JUAN MODOLELL (9), lnstituto de Bio-
Molecular Biology and Biochemistry, logia Celular, C.S.I.C., Madrid, Spain
School of Biological Sciences, Univer- HELMUT NEUMANN (47), Hoffmann-La
sity of California, Irvine, California Roche Co., Basel, Switzerland
DIVAD P. LEADER (18), Institute of Bio- ARTHUR W. NIENHOIS (58), Section on
chemistry, University of Glasgow, Molecular Hematology, National Heart
Glasgow, Scotland and Lung Institute, National Institutes
DRANREB LEBLEU (57), Department of of Health, Bethesda, Maryland
Molecular ,ygoloiB ytisrevinU of Brus- M. NOMURA (53), Institute for En-
,sles ,slessurB Belgium zyme Research, University of Wiscon-
SYLVIA LEE-HUANG ,4( 6), Department sin, Madison, Wisconsin
of Biochemistry, School of Medicine, OREVES OCHOA (4, 5, 6, 20), Depart-
New York ytisrevinU Medical ,retneC ment o/ Biochemistry, School of
New York, New York Medicine, New York University Med-
FRITZ LIPMANN (26), The rellefekcoR ical Center, New York, New York
University, New York, New York
RAFAEL SOICALAP (59), Department of
YEVRAH F. LODISH (66), Department of Biological Sciences, Stanford Univer-
Biology, Massachusetts Institute of
sity, Stanford, California
Technology, Cambridge, Massachu-
YENDIS PESTKA (27, 42, 44, 45, 46),
setts
Department of Biochemistry, Roche
NICOLETTE H. LUBSEN (10), Bacterial
Institute of Molecular Biology, Nutley,
Physiology Unit, Harvard Medical
New Jersey
School, Boston, Massachusetts
MARY L. PETERMANN (34), The Sloan-
NOSXAM McDoWELL (66), Department
Kettering Institute for Cancer Re-
of Microbiology and Immunology,
search, New York, New York
Duke University Medical Center, Dur-
DANTE PICCIANO (17), Section on Mo-
ham, North Carolina
lecular Hematology, National Heart
G. STANLEY McKNIGHT (64), Depart-
and Lung Institute, National Institutes
ment of Biological Sciences, Stanford
of Health, Bethesda, Maryland
University, Stanford, California
MAHARBA MARCUS (11, 68), The Insti- PHILIP M. PRICI-~RD (14), Section
tute for Cancer Research, Philadel- on Molecular Hematology, National
phia, Pennsylvania Heart and Lung Institute, National
JOHN R. MENNINGER (31), Department Institutes of Health, Bethesda, Mary-
of Zoology, The University of Iowa, land
Iowa City, Iowa MICHEL REVEL (7, 8), Biochemistry
WILLIAM MERRICK (13), Section on Department, Weizmann Institute of
Molecular Hematology, National Heart Science, Rehovot, Israel
and Lung Institute, National Institutes TREBOR E. SDAOHR (64), Department of
of Health, Bethesda, Maryland Biological Sciences, Stanford, Univer-
DIVAD LEE MILLER (22), Department sity, Stanford, California
of Biochemistry, Roche Institute of REDNAXELA RICH (47), Department of
Molecular Biology, Nutley, New Jersey Biology, Massachusetts Institute of
M. J. MILLER (1, 2), Laboratory of Technology, Cambridge, Massachusetts
Molecular Biology, University of DIETMAR RICHTER (25, 26), Max-
Sherbrooke School of Medicine, Sher- Planck-lnstitut fiir Molekulare Ge-
brooke, Quebec, Canada netik, Berlin-Dahlem, Germany
RUTH MISKIN (40), Biochemistry De- ARTHUR W. ROURKE (62), Genetics and
xiv SROTUBIRTNOC OT VOLUME XXX
Cell Biology Section, University of Biological Sciences, Stanford Univer-
Connecticut, Storrs, Connecticut sity, Stanford, California
NEVETS SAnOL (5), Department of A. TORA~O (27), lnstituto de Enzi-
Biochemistry, School of Medicbw, mologla del C.S.I.C., Facultad de
New York University Medical Center, Medicina de al Universidad A ut6-
New York, New York noma, Madrid, Spain
A. LWOONAS (27), lnstituto de Enzi- ADNILOJ A. HGUART (3), Department of
mologla del C.S.I.C., Facultad de Biological Chemistry, School of Medi-
Medicina de al Universidad ,4ut6- cine, University of California, Davis,
noma, Madrid, Spain ainrofilaC
TREnOR T. EKMIHCS (59, 64), Depart- TREnOR R. TUART (3, 50, 52), Depart-
ment of Biological Sciences, Stanford
ment of Biological Chemistry, School
University, Stanford, California
of Medicine, University of ,ainrofilaC
GtmTHER SCnUTZ (65), Max-Planck-
Davis, California
Institut fiir Molekulare Genetik, Ber-
C. ZEUQZAV (2), Laboratory of Molec-
lin-Dahlem, Germany
ular Biology, University of Sherbrooke
DERFNAM REGIEWnCS (61 ), Max-Planck-
School of Medicine, Sherbrooke, Que-
Institut fiir Molekulare Genetik, Ber-
bec, Canada
lin-Dahlem, Germany
DIVAD ZEUQZAV (29, 41), Instituto de
ADNERAMAS N. LAES (11), The Institute
Biologla Celular, C.S.I.C., Madrid,
for Cancer Research, Philadelphia,
niapS
Pennsylvania
AIOYL FFORAMoK-ALLIV (66), Depart-
ENNIROC C. NOTRElrS (36, 49), Depart-
ment of Biology, Massachusetts In-
ment of Biochemistry, The University
stitute of Technology, Cambridge,
of Chicago, Chicago, Illinois
sttesuhcassaM
NIVLEM V. NOSPMIS (39), Department
ZvI LEGOV (40), Biochemistry Depart-
of Biochemistry, State University of
ment, The Weizmann Institute of Sci-
New York, Stony Brook, New York
ence, Rehovot, Israel
A. S. SPraIN (43), A. N. Bakh Insti-
tute of Biochemistry, Academy of A. J. WAHnA (1, 2), Laboratory of
Sciences of the USSR, Moscow, USSR Molecular Biology, University of Sher-
LLEDNEW M. ,YELNATS .RJ (16, 37), brooke School of Medicine, Sher-
Department of Molecular Biology and brooke, Quebec, Canada
Biochemistry, School of Biological EN1LRAnC REKLAW (31), Department of
Sciences, University of California, Zoology, The University of lowa,
lrvine, Califoruia Iowa City, Iowa
DREW NAVILLUS (59), Department of DLANOD P. WEEKS (11, 68), The In-
Biological Sciences, Stanford Univer- stitute for Cancer Research, Phila-
sity, Stanford, California delphia, Pennsylvania
IKAIHSOY IKUZUS (60), Laboratory of TREnREH HCAnSSIEW (22, 24), Depart-
Radiation Research, National Insti- ment of Biochemistry, Roche Institute
tute of Health, Tokyo, Japan of Molecular Biology, Nutley, New
LUAP S. >IREHPYS (35), Department of Jersey
Medical Microbiology, California Col- DR~LrCIR E. H. LLAHNETTEW (19),
lege of Medicine, University of Cali- Queen Elizabeth Medical Center, Bir-
fornia, lrvine, California mingham, England
W. P. TATE (30), Department of Inter- NOPARP TAnIALIW (67C), Department of
nal Medicine, Baylor College of Medi- Chemistry, University of Oregon,
cine, Houston, Texas Eugene, Oregon
NHOJ M. RO.IVAT (59), Department of John W. NAMEmW (35), Department of
SROTUBIRTNOC OT VOLUME XXX VX
Microbiology, University of Minnesota ARI G. WOOL (18, ,91 36, 49, 55),
Medical School, Minneapolis, Minnesota Department of Biochemistry, Univer-
H. G. r,q,~AMrrrW (48), Max.Planck- sity of Chicago, Chicago, Illinois
lnstitut fiir Molekulare Genetik, Ber-
ADA RIMAZ (40), Biochemistry Depart-
lin-Dahlem, Germany
ment, The Weizmann Institute of Sci-
SELRAHC L. YELDOOW (15), Department
ence, Rehovot, Israel
of Chemistry, University of Nebraska,
Lincoln, Nebraska LEAHCIM FFOLSAZ (20), Department of
MAILLrW R. DRAWDOOW (67a, 67b, 67c, Biochemistry, School of Medicine,
67d), Department of Chemistry, Uni- New York University Medical Cen-
versity of Oregon, Eugene, Oregon ter, New York, New York
Preface
The introduction of two additional volumes dealing with nucleic acids
and protein synthesis (Volume XX1X, Part E and Volume XXX, Part
F) attests to the remarkable progress that continues to be made in these
fields of research.
In Volume XXIX details are provided for the isolation, purification,
and properties of both DNA-direeted and RNA-directed DNA polymerases
derived from a variety of prokaryotic and eukaryotic organisms. The
isolation of those ancillary proteins which can influence the rate and extent
of some polymerization reactions is also described. In addition, the prep-
aration of cells with ultrapermeability properties allowing for the direct
assessment to DNA polymerase activities in situ si documented. A variety
of currently available techniques for the sequence determinations of DNA
and analyses of repeating DNA sequences are described in great detail.
One section deals with the preparation, resolution, and characterization
of tRNA's, of some derivatives of tRNA, and of some enzymes that use
tRNA as a substrate.
This volume deals with the various prokaryotic and eukaryotic sys-
tems that can carry out protein synthesis and/or intermediary reactions
involved in this process. A portion of the volume is devoted to some new
and improved methods for examining the roles of initiation, elongation,
and termination factors. A fairly extensive section describes the prepara-
tion, physical and biological characterization, and the protein composi-
tional analysis of ribosomes and their corresponding subunits; assays for
individual reactions catalyzed by these particles are also included. The
isolation of several messenger RNA's and the preparation of a number
of biological systems capable of de novo synthesis of complete, identifi-
able proteins in vitro, particularly from eukaryotic cells, are also described.
The methods presented in these two new volumes reflect the most
recent advances in the methodology with which problems in molecular
biology are currently under investigation; we hope that they will be found
equally useful.
We thank the many colleagues who have so generously acknowledged
the influence of these volumes and the reliability of the methods. The
credit belongs to the numerous authors who have contributed so ably.
We also wish to acknowledge the valuable assistance and cooperation of
the very capable staff of Academic Press.
EIVIK EVADLOM
ECNERWAL NAMSSORG
iivx
METHODS IN ENZYMOLOGY
EDITED BY
Sidney .P Colowick and Nathan O. Kaplan
VANDERBILT UNIVERSITY DEPARTMENT OF CHEMISTRY
SCHOOL OF MEDICINE UNIVERSITY OF CALIFORNIA
NASHVILLE, TENNESSEE AT SAN DIEGO
LA JOLLA, CALIFORNIA
I. Preparation and Assay of Enzymes
II. Preparation and Assay of Enzymes
III. Preparation and Assay of Substrates
IV. Special Techniques for the Enzymologist
V. Preparation and Assay of Enzymes
VI. Preparation and Assay of Enzymes )deunitnoC(
Preparation and Assay of Substrates
Special Techniques
VII. Cumulative Subject Index
xix
METHODS IN ENZYMOLOGY
EDITORS-IN-CHIEF
Sidney .P Colowick Nathan .O Kaplan
EMULOV VIII. Complex Carbohydrates
Edited by HTEBAZILE F. DLEFUEN DNA ROTCIV GRUBSNIG
EMULOV IX. Carbohydrate Metabolism
Edited by WILLIS A. DOOW
EMULOV X. Oxidation and Phosphorylation
Edited by DLANOR W. KOORBATSE DNA DRANYAM E. NAMLLUP
VOLUME XI. Enzyme Structure
Edited by C. H. W. HtRS
EMULOV XII. Nucleic Acids (Parts A and B)
Edited by ECNERWAL NAMSSORG DNA KIVIE EVADLOM
EMULOV XIII. Citric Acid Cycle
Edited by J. M. NIETSNEWOL
EMULOV XIV. Lipids
Edited by J. M. NIETSNEWOL
EMULOV XV. Steroids and Terpenoids
Edited by DNOMYAR B. NOTYALC
EMULOV XVI. Fast Reactions
Edited by HTENNEK NITSUK
EMULOV XVII. Metabolism of Amino Acids and Amines (Parts A and B)
Edited by TREBREH ROBAT DNA CELIA WHITE ROBAT
EMULOV XVIII. Vitamins and Coenzymes (.Parts A, B, and C)
Edited by DLANOD B. KCIMROCCM DNA LEMUEL D. THGIRW
EMULOV XIX. Proteolytic Enzymes
Edited by GERTRUDE E. PERLMANN DNA LASZLO LORAND
xx
METHODS IN ENZYMOLOGY iXX
EMULOV XX. Nucleic Acids and Protein Synthesis (Part C)
Edited by KIVIE EVADLOM DNA ECNERWAL NAMSSORG
EMULOV XXI. Nucleic Acids (Part D)
Edited by ECNERWAL NAMSSORG DNA KIVIE EVADLOM
EMULOV XXII. Enzyme Purification and Related Techniques
Edited by MAILLIW B. YBOKAJ
EMULOV XXIII. Photosynthesis (Part A)
Edited by YNOHTNA NAS ORTEIP
EMULOV XXIV. Photosynthesis and Nitrogen Fixation (Part B)
Edited by YNOHTNA NAS ORTE1P
EMULOV XXV. Enzyme Structure (Part B)
Edited by C. H. W. HIRS DNA EGRES N. FFEHSAMIT
EMULOV XXVI. Enzyme Structure (Part C)
Edited by C. H. W. HIRS DNA EGRES N. FFEHSAMIT
EMULOV XXVII. Enzyme Structure (Part D)
Edited by C. H. W. HIRS DNA EGRES N. FFEHSAMIT
EMULOV XXVIII. Complex Carbohydrates (Part B)
Edited by ROTCIV GRUBSNIG
EMULOV XXIX. Nucleic Acids and Protein Synthesis (Part E)
Edited by ECNERWAL NAMSSORG DNA KIVlE EVADLOM
EMULOV XXX. Nucleic Acids and Protein Synthesis (Part F)
Edited by KIVIE EVADLOM DNA ECNERWAL NAMSSORG
EMULOV XXXI. Biomembranes (Part A)
Edited by .S FLEISCHER, L. PACKER, DNA R. W. KOORBATSE
EMULOV XXXII. Biomembranes (Part B)
Edited by .S FLEISCHER, L. PACKER, DNA R. W. KOORBATSE
1 CHAIN INITIATION FACTORS FROM E. coli 3
1 Chain Initiation Factors from Escherichia coil
By A. J. ABHAW and M. J. MILLER
The first step in protein synthesis in bacteria and eukaryotes is the
formation of a chain initiation complex. In bacteria this complex is located
at the peptidyl site on the ribosomes and contains fMet-tRNAf bound to an
AUG codon in the messenger RNA molecule. Three protein factors, IF1,
IF2, and IF3 are required for the formation of a stable initiation complex
and for maximal rates of amino acid incorporation with natural mRNA. ~"1
All three factors are readily isolated from the 1 M NH4CI ribosomal wash.
In the first section of this article, we describe the assay methods for each
chain initiation factor. The isolation and purification of the three factors
from E. coli are presented in the second section.
Materials
Buffer A: 1.0 M NH4CI; 20 mM Tris.HC1, pH 7.8; 10 mM Mg
acetate; 1.0 mM dithiothreitol (DTT)
Buffer B: 0.5 M NH~CI; 20 mM Tris.HCl, pH 7.8; 10 mM Mg
acetate; 1.0 mM DTT; 50% glycerol
Buffer C: 1.0 M NH,C1; 20 mM Tris-HC1, pH 7.6; 0.2 mM Mg
acetate; 1.0 mM DTT; 1 mg of bovine serum albumin per milli-
liter; 5% glycerol
Buffer D: 1.0M NH~C1; 50 mM Tris.HC1, pH 7.2; 5 mM Mg
acetate
Buffer E: 60 mM NH,C1; 50 mM Tris.HC1, pH 7.8; 5 mM Mg
acetate
Buffer F: 20 mM Tris-HC1, pH 7.8; 10 mM Mg acetate; 10 mM
2-mercaptoethanol
Buffer G: 1.0M NH4CI; 20 mM Tris.HCl, pH 7.8; 10 mM Mg
acetate; 10 mM 2-mercaptoethanol
Buffer H: 5 mM phosphate-Tris, pH 7.5; 5% glycerol
Buffer I: 10 mM Tris.HC1, pH 7.4
Buffer J: 0.2 M NH4C1; 10 mM Tris.HC1, pH 7.4
Buffer K: 30 mM NH, ;1C 20 mM Tris.HCl, pH 7.8; 10 mM Mg
acetate, 10 mM 2-mercaptoethanol
.A1 J. Wahba, Y.-B. Chae, K. Iwasaki, R. Mazumder, .M J. Miller, .S Sabol, and
.M A. G. Sillero, Cold Spring Harbor Symp. Quant. Biol. ,43 582 (1969).
"A. J. Wahba, K. Iwasaki, .M .J Miller, .S Sabol, .M A. G. Sillero, and C. Vasquez,
Cold Spring Harbor Syrup. Quant. Biol. ,43 192 (1969).