Nucleic Acids, Part B

962 Pages·1968·26.922 MB·English

by  Lawrence Grossman, Kivie Moldave (Eds.)

Contributors to Volume XII, Part B
Article numbers are shown in parentheses following the names of contributors.
AiHliations listed are current.
EGROEG Acs (149), Microbiology Institute ment o Biochemistry, University o
o/ Muscle Disease, New York, New Kentucky Medical School, Lexington,
York Kentucky
ECILA J. R.~LDA (122), Graduate Depart- ELLIS T. NOTLOB (147), Department o
ment o Biochemistry, Brandeis Uni- Terrestrial Magnetism, Carnegie Insti-
versity, Waltham, Massachusetts tution o Washington, Washington,
H. VAS~N NAIHSOPA (142), Department D.C.
o Microbiology, TuIts University SEMAJ RENNOB (96), Division o Biology,
School o Medicine, Boston, Massa- Caliornia Institute o Technology,
chusetts Pasadena, Caliornia
RALP~ SU~L~ONI~rA (153), United States M. S. ,FHCSTERB a (164), MRC Labora-
Department o Agriculture, Animal tory o Molecular Biology, Cambridge,
Disease and Parasite Research Divi- England
sion, Plum Island Animal Disease D. P. BURMA (141b), Department o
Laboratory, Greenport, New York Biochemistry and Biophysics, Banaras
J. T. Auovsv (137), Department o Hindu University, Varanasi, India
Molecular Biology, Albert Einstein K. NOTRUB (105), Department o Bio-
College o Medicine, Yeshiva Univer- chemistry, University o Oxord, Ox-
sity, Bronx, New York lord, England
LEAHCIM REEB (126), Biophysics Depart- SIRRAH BUSCH (97), Department o
ment, The Johns Hopkins University, Pharmacology, Baylar University Col-
Baltimore, Maryland lege o Medicine, Texas Medical Cen-
MAS M. RESIEB (173), Department o ter, Houston, Texas
Microbiology, College o Physicians P. CERV~rI (130), Department o Chem-
and Surgeons, Columbia University, istry, Princeton University, Princeton,
New York, New York New Jersey
A. R. YMALLEB (104), Department o G. REGOR YELKLAHC (96), Department
Cell Biology, Albert Einstein College o Biochemistry, University o Iowa
o Medicine, Yeshiva University, Medical School, Iowa City, Iowa
Bronx, New York HPESOJ D. LIYAREHC (106a), Indian In-
DLONRA J. BENI)ICH (147), Department stitute o Science, Bangalore, India
Jo Microbiology, University o Wash- R. A. Cox (103a), Medical Research
ington, Seattle, Washington Council, National Institute or Med-
ARABRAB REKAROJB (116b), Molecular ical Research, London, England
Biology Laboratory, University o LEAHCIM SUMHAD (96), Division o Biol-
Wisconsin, Madison, Wisconsin ogy, CaIiornia Institute o Technol-
R. M. BocK (106a, 116b, 120), Molecular ogy, Pasadena, Caliornia
Biology Laboratory, University o TELOIV DANIEL (141C), Biochemistry
Wisconsin, Madison, Wisconsin Section, The Weizmann Institute o
AGLEH REKrYEOB (129), Biology Depart- Science, Rehovoth, Israel
ment, Harvard University, Boston, H. DELIUS (172), Laboratoire de Bio-
Massachusetts chimie, Universitd de Gen$ve, Geneva,
F. J. MULLOB (106b, 143, 144), Depart- Switzerland
vi SROTUBIRTNOC OT VOLUME XII, PART B
TREBREH W. NAMREKCID (150b), The o Microbiology, Tuts University
Johns Hopkins University, Baltimore, School o Medicine, Boston, Massa-
Maryland chusetts
HPESOJ EmNER (128), Department o GNIVRI H. GREBDLOG (135), Beth Israel
Microbiology, Washington University Hospital, Boston, Massachusetts
School o Medicine, St. Louis, Missouri LoS H. LAGBOOG (170), Department o
S. RETLAW REDNALGr~E (127), Biochem- Microbiology, University o Pennsyb
istry Department, University o Penn- Bahia School o Medicine, Philadel-
sylvania School o Medicine, Phila- phia, Pennsylvania
delphia, Pennsylvania Faa~c~ois Gaos (158), Institut de Biolo-
Na.rLL1L GNAYOE (137), Department o cig Physico-Chimique, Service de
Molecular Biology, Albert Einstein Physiologic Microkienne, Paris, France
College of Medicine, Yeshiva Univer- ECNERWAL NAMSSORG (131), Graduate
sity, Bronx, New York Department o Biochemistry, Brandeis
)mXNaEB F. REGNatRE (173), Department University, Waltham, Massachusetts
o Microbiology, College o Physicians M. OGANAM-GREBNURG (136), Institut
and Surgeons, Columbia University, de Biologic Physico-Chimique Foun-
New York, New York dation Edmond de Rothschild, Paris.
Douot~s HGUORBMAF (96), Division o ecnarF
Biology, Caliornia Institute of Tech- DLONRA ItAMPEL (106a), Molecular Bi-
nology, Pasadena, California ology Laboratory, University o WIS-
G~B D. FAS~AN (122), Graduate De- eonsin, Madison, Wisconsin
partment o Biochemistry, Brandeis I. H~u~A (138), Department o Molec-
University, Waltham, Massachusetts ular Biology School o/Medicine, Keio
YRAG VLEFNESLEF (119), Department o University, Shinjuku-Ku, Tokyo,
Health, Education and Welare, Na- Japan
tional Institutes o Health, Bethesda, Do~r~ H. SEYAH (158), Institut de Biol-
Maryland ogic Physlco-Chimique, Service de
H. FR~N~L-CoNRAT (114, 118), Depart- Physiologic Microkienne, Paris, France
ment o Molecular Biology, University REOOR I~I~TZ (153), Biochemistry De-
o CaliIornia at Berkeley, Berkeley, partment, University o Kentucky
CaliIornla School o Medicine, Lexington, Ken-
DRAHC~I M. F~rKta~ (141a), Depart- tucky
ment o Virology, The Public Health r~oEL A. H~PPSr. (166), CorneU Univer-
Research Institute o the City o New sity, Ithaca, New York
York, Inc., New York, New York J. L. HOr~M^N (120), Department o
J. D. FRI~.SEN (146), Department o Biochemistry, University o Louisville,
Physics, Kansas State University, Louisville, Kentucky
Manhattan, Kansas P. H. REDIENHCSFOI'I (172), Max-Planck-
FRANK FUJIMURA (96), Department o/ Institut iJr Biochemie, Mi2nehen,
Biochemistry and Molecular Biology, Germany
Harvard University, Cambridge, Mas- WmL~M J. YELsRoHH (140), Department
sachusetts o Chemistry, University o ,ainroilaC
D. mPSELLIG (148), Biology Department, Berkeley, Caliornia
Brandeis University, Waltham, Massa- RU-CHIH C. GN^uHH (96), Department o/
chusetts Biology, The Johns Hopkins Univer-
Mnaw~¢ GoLD (132, 133a), Medical -OIB sity, Baltimore, Maryland
physics, University oI Toronto, LEOJ NAMREBUH (96), Department o/
Toronto, Canada Biochemistry, Stanord University
Eaw^aa B. G~BDLOG (169), Department Medical School, Palo Alto, California
SROTUBIRTNOC OT VOLUME XII~ PART B vii
J. A. HUNT (117), Department o Genet- NEHPETS H. ALPPEL (l16b), Molecular
ics, University o Hawaii, Honolulu, Biology Laboratory, University o Wis-
Hawaii consin, Madison, Wisconsin
J. ZTIWRUH (110, 132, 133a), Department ZTIRF LneMA~N (149, 154), The Roeke-
o Molecular Biology, Albert Einstein letter University, New York, New York
College o Medicine, Yeshiva Univer- LmRU Z. LI~VA~R (134, 141e), Biochem-
sity, Bronx, New York istry Section, The Welzmann Institute
DLA~OR JENSEN (96), Growth Sciences o Science, Rehovoth, Israel
Center, International Minerals and JEAN LucAs-LE~ (154), The Rocke-
Chemical Corporation, Libertyville, eller University, New York, New
Illinois York
JOHN JOSSE (133b), Syntex Institute o B. J. McCARTHY (163a,b), Department
Molecular Biology, Pals Alto, Cali- o Microbiology, University o Wash-
ainro ington School o Medicine, Seattle,
A. D. KAISER (171), Biochemistry De- Washington
partment, StanIord University School AGNI RELI.IAM (167), Graduate Depart-
o Medicine, Pals Alto, CaliIornia ment o Biochemistry, Brandeis Uni-
AKII~ IZ~jI (152), Department o Micro- versity, Waltham, Massachusetts
biology, University o Pennsylvania (cid:127). AINKNELAM (157b), Laboratoire de
School o Medicine, Philadelphia, Recherches de Biochimie Medicale,
Pennsylvania H~pital des Enants-Malades, Paris,
FESOY KIMHI (134), Biology Division, France
Massachusetts Institute o Technol-
M. LEDNAM (108, 109), Department o
ogy, Cambridge, Massachusetts
Biology, The University o Texas, M.
K. S. KIRBy (98), Institute o Cancer
D. Anderson Hospital and Tumor In-
Research, Chester Beatty Research In-
stitute, Houston, Texas
stitute, Pollards Wood Research Sta-
J. M~M~ (108, 109), Biochemistry De-
tion, London, England
partment, Albert Einstein College o
A. K. T)nMHCSNXELK (125), Department
Medicine, Yeshiva University, Bronx,
o Biochemistry, New York University
New York
School o Medicine, New York, New
KEIJI EGIHSURAM (96), Department o
York
Biochemistry, University o British
TREBOR P. TTELK (101), The RockeIeller
Columbia, Vancouver, British Colum-
University, New York, New York
bia, Canada
HPESOJ S. KI~Kow (140), Space Sciences
r~oTLA MEISTga (150a), Department o
Laboratory, University o Caliornia
Biochemistry, CorneU University
at Berkeley, Berkeley, CaliIornia
School o Medicine, New York, New
SEUQCAJ KaUH (157a), Laboratoire de
York
Recherches de Bioehimie Medicale,
HSpital des Enants-Malades, Paris, H. DDOT MILES (121), National Insti-
France tutes o Health, Bethesda, Maryland
F. EVOCAL (176), Institut Gustav Roussy, Klvm WADLOM (155), Department o
Villejui, France Biochemistry, University o Pittsburgh
CY~THt~ LARK (145), Department o School o Medicine, Pittsburgh, Penn-
Physics, Kansas State University, sylvania
Manhattan, Kansas WILFRma F. H. M. STEEA~MOM (123),
PHmIP aEDEL (156, 165a), Protein Chem- Department o Physiology, University
istry Section, Laboratory o Biochem- o Caliornia School o Medicine, The
istry, Department o Health, Education Center or the Health Sciences, Los
and Welare, National Cancer Ir~ti- Angeles, Caliornia
tute, Bethesda, Maryland NALLA J. MoaaIS (165), Department o
Vlll .°° CONTRIBUTORS OT VOLUME XII~ PART B
Biochemistry, Michigan State Univer- School o Medicine, Boston, Massa-
sity, East Lansing, Michigan chusetts
.TmNAD SNAHTAN (160), Department o DRAHCIR *TEEWHCS (153), Department
Microbiology, The Johns Hopkins Uni- o Cell Biology, University o Ken-
versity School o Medicine, Baltimore, tucky Medical Center, Lexington,
Maryland Kentucky
IMOTUSAY NISHIZUKA (154), Department ZSSEJ F. TTOCS (106e), The John Collins
o Medical Chemistry, Kyoto Univer- Warren Laboratories, Massachusetts
sity Faculty o Medicine, Kyoto, Japan General Hospital, Boston, Massachu-
B. NISMAN (162), Laboratoire d'Enzy- setts
mologie Microbienne, CNRS, Gi/-sur- ANDE SE^Mn~ (112), Graduate Depart-
Yvette, France ment o Biochemistry, Brandeis Uni-
HANS LLON (103b), Department o Bio- versity, Waltham, Massachusetts
logical Sciences, ~ Northwestern Uni- LEAHCIM SELA (175), The Weizmann
versity, Evanston, Illinois Institute ol Science, Rehovoth, Israel
A. YESDORGOVON (110), Department o TREBOR L. SINSHEIMER (168), Division
Biophysics, The Weizmann Institute o Biology, Cali/ornia Institute o/
o Science, Rehovoth, Israel Technology, Pasadena, Caliornia
I:~IKO HCSUBNELHO (96), Departement LEAHCIM SMITH (101), Department o
de Bioehimie Macromoleculaire, Centre Biochemistry, Faculty o Medicine,
National de al Recherche Scientifique, University o British Columbia, Van-
Montpellier, France couver, British Columbia, Canada
OREMODLAB AREVILO (96), Department o S. NAMLEGEIPS (138), Departznent o
Biochemistry, Stan/ord University Microbiology, University o Illinois,
Medical School, Palo Alto, Caliornia Urbana, Illinois
N. R. PAcE (138), Department o Micro- A. REDIENHCSNIETS (118), Hebrew Uni-
biology, University o Illinois, Urbana, versity o Jerusalem, Department o
Illinois Biological Chemistry, Jerusalem, Israel
TREBREH STERN (100), Department o
P. UmDAP (157b), Laboratoire de Recher-
Botany, University o Illinois, Urbana,
ches de Biochimie Medicale, H~pital
Illinois
des Enants-Malades, Paris, France
AuDa~r SNEVETS (161), Biology Division,
Orro J. ~LCSELP (174), Institute O Mi-
Oak Ridge National Laboratory, Oak
crobiology, Rutgers University, New
Ridge, Tennessee
Brunswick, New Jersey
DEAHRE SrUTZ (103b), Department o
R. K. RALPH (104), Microbiology De-
Biological Sciences, Northwestern Uni-
partment, Auckland University, Auck-
versity, Evanston, Illinois
land, New Zealand
WALCAW k'kSLASVZS (124), McArdle Lab-
J. Rosa (157b), Laboratoire de Recher- oratory or Cancer Research, Univer-
ches de Biochimie Medicale, H~pital
sity o Wisconsin, Madison, Wiscon-
des Enants-Malades, Paris, France sin
G. AmPAHCS (157b), Centre Universitaire I. TAXAHASHI (99), Biochemistry Re-
Port Royal, Cochin-lnstitut de Pathol- search Unit, McMaster University,
ogle Moleculaire, Pariz, France Hamilton, Ontario
C. L. TUARKDLIHCS (108), Department o TRAUTS W. MUABNENAT (173), Depart-
Cell Biology, Albert Einstein College ment o Microbiology, College o Phy-
o Medicine, Yeshiva University, sicions and Surgeons, Columbia Uni-
Bronx, New York versity, New York, New York
GERHARD SCHMII)T (l16a), Department
o Biochemistry, Tuts University * Deceased April 3, .7691
CONTRIBUTORS OT VOLUME XlI~ PART B ix
G. M. TENER (113), Department o Bio- Biology, State University o New York
chemistry, University o British Co- at Bulalo, Bualo, New York
lumbia, Vancouver, British Columbia, I. DRANREB WEINSTEIN (159), Institute
Canada o Cancer Research, College o Physi-
M. N. TH^NG (136), Institut de Biologie cians and Surgeons, New York, New
Physico-Chimique, Biochemistry Divi- York
sion, Paris, France LEUMAS B. WEISS (139), Argonne Cancer
A. Toa~NI (111), Department o Bi- Research Hospital, University o
ology, Massachusetts Institute o Tech- Chicago, Chicago, Illinois
nology, Cambridge, Massachusetts VREBaEH WEISSBACH (150b), National
ANNAH NoRAW-RAGNU (175), The Weiz- Heart Institute, National Institutes o
mann Institute o Science, Rehovoth, Health, Bethesda, Maryland
Israel JACK WIDHO~ (96), Growth Sciences
GF~IT NESIUHDLEV (169), Medical Bio- Center, International Minerals and
logical Laboratory o the National De- Chemical Corporation, LibertyviUe,
lense Research Council TNO, Rijs- Illinois
wijk, Z. H. Netherlands DRAHCIR NEDNEFLOW (151), Frick Chemi-
SHIH Y1 WANG (107), Department o cal Laboratory, Princeton University,
Biochemistry, The Johns Hopkins Uni- Princeton, New Jersey
versity School o Public Health and YERFFOEG L. YABUZ (115), Department
Hygiene, Baltimore, Maryland o Zoology, Columbia University, New
TUNG EUY WANG (102), Department o York, New York
Preface
Part A of this work si devoted to methods and techniques involved
with the chemistry and enzymology of nucleic acid components. It also
emphasizes the preparation of oligonucleotides and their subsequent
resolution by eleetrophoresis and chromatography. The preparation of
tissue organelles for the isolation and fractionation of nucleic acids si
also described.
This volume, Part B, deals with the characterization of isolated and
resolved DNA and RNA by base content, concentration, and by chemi-
cally and enzymatically deduced chain-length measurements. Conforma-
tion and molecular weight assignments, by optical, physical, and isotopic
exchange techniques, and end-group modification procedures are also
presented. A section of this volume concentrates on the specific interaction
of nucleic acids: with enzymes involved in modifications through methyla-
tion, glycosylation, and aminoacylation of tRNA; with similar nucleic
acids capable of hybridization; and in directing replication and transcrip-
tion. The enzymatic techniques incident to sequential base specificities
in amino acid coding and protein synthesis are presented. The conforma-
tional limitations of nucleic acids with respect to interactions in biological
systems are considered, as well as the specific sequential and conforma-
tional requirements for the interaction of nucleic acids with their
homologous antibodies.
The current progress in describing the subtle sequential and conforma-
tional limitations of proteins is being realized today, allowing an under-
standing of the specificity of enzymes. Such knowledge, with respect to
nucleic acids, si forthcoming; it is the hope of the editors that these
volumes will assist the scientific community to achieve a similar goal,
with respect to an understanding of the active sites within nucleic acids,
by providing a compilation of pertinent techniques.
The editors wish to thank the contributors for their cooperation and
their valuable contributions to the completion and ultimate usefulness
of these volumes, and Barbara Grossman and Rose Moldave for their
patience and understanding. We would also like to thank the staff of
Academic Press for their cooperation during the production of this work.
March, 8691
ECNERWAL NAMSSOR~
EIVIK EVADLOM
ix
METHODS IN ENZYMOLOGY
DETIDE YB
Sidney .P Colowick dna Nathan .O nalpaK
VANDERBILT UNIVERSITY GRADUATE DEPARTMENT OF
SCHOOL OF MEDICINE BIOCHEMISTRY
NASHVILLE 3 TENNESSEE BRANDEIS UNIVERSITY
WALTHAM~ MASSACHUSETTS
I. Preparation and Assay of Enzymes
II. Preparation and Assay of Enzymes
III. Preparation and Assay of Substrates
IV. Special for Techniques the Enzymologist
.V Preparation and Assay of Enzymes
.IV Preparation and Assay of Enzymes }deunitnoC(
Preparation and Assay of Substrates
Techniques Special
VII. Cumulative Subject Index
METHODS IN ENZYMOLOGY
EDITORS-IN-CHIEF
yendiS P. Colowick Nathan .O nalpaK
EMULOV VIII. Complex Carbohydrates
by Edited ttTEBAZILE F. DLEFUEN DNA ROTCIV GRUBSNIG
EMULOV IX. Carbohydrate Metabolism
by Edited SILLIW .A DOOW
EMULOV .X Oxidation and Phosphorylation
Edited by RONALD W. ESTABROOK DNA DRANYAI(cid:127) E. PULLMAN
EMULOV XI. Enzyme Structure
Edited by C. H. W. Hms
EMULOV XII. Nucleic Acids ( in two volumes )
Edited by ECNERWAL NAMSSORG DNA EIVIK EVADLOM
In noitaraperP
Citric Acid Cycle
Edited by .J .M NIETSNEWOL
Lipids
by Edited .J .M NIenTSNEWOL
Steroids and Terpenoids
Edited by DNOMYAR .B NOTYALC
Fast Reactions
Edited by HTENNEK NITSUK
Metabolism of Amino Acids and Amines
Edited by TREBREH ROBAT DNA AILEC ETIHW RORAT
Vitamins and Coenzymes
Edited by DONALD B. ~icCoRM1CK DNA LEMUEL D. WRIGHT
Proteolytic Enzymes
by Edited GERTRUDE E. PERLMANN DNA LASZLO LORAND
Photosynthesis
Edited by .A NAS OaTmP
Enzyme Purification and Related Techniques
Edited by WmraAM .B YBOKAJ
Errata for Volume XII, Part A
p. vi. The affiliation for John W. Little should read "National Institute of Arthritis
and Metabolic Diseases, National Institutes of Health, Bethesda, Maryland."
p. vii. The affiliation for B. Setlow should read "Graduate Department of Bio-
chemistry."
p. 164. Footnote 6 should read "B. Setlow" instead of "R. B. Setlow."
p. 169. Footnote 11 should read "B. Setlow" instead of "R. B. Setlow."
p. 722. Entry in author index: Setlow, B., 164, .961
The following footnote should be added to Articles 4 and 26, pp. 23 and 214,
respectively :
Refluxing 100% hydrazine hydrate with an equal weight of sodium hydroxide
pellets for 2 hours and then distillation in a slow stream of nitrogen introduced
through a capillary leak is recommended. Distillation in air can lead to an
explosion.
69 LAMOSOMORHC SNIETORPOELCUN 3
96 Isolation and Characterization of
Chromosomal Nucleoproteins
By SEMAJ ,RENNOB G. REGOR ,YELKLAHC MICHAEL ,SUMHAD
SALGUOD ,HGUORBMAF FRANK FUJIMURA, RU-CHIH G. ,GNAUH
LEOJ ,NAMREBUH DLANOR JENSEN, KEIJI ,EGIHSURAM HEIKO
,HCSUBNELHO OREMODLAB ,AREVILO and JACK MLOHDIW
I. Introduction and Scope
This chapter considers the isolation and characterization of chromo-
somes. Chromosomes are ordinarily obtained from cells during inter-
phase and are therefore in the extended form known as chromatin. Isola-
tion of the highly contracted metaphase chromosomes is also possible,
however, and is also dealt with below. The recent rapid advances in our
understanding of chromosomal structure and function have been made
possible by the development of new methods for the handling of chromatin
and of chromosomal constituents. The methods described below are all
ones which the authors have used, and which have proven satisfactory
for their several purposes.
II. Isolation of Chromatin
Principles
The isolation of chromatin is based upon differential centrifugation
followed by sucrose density gradient centrifugation. 1 Chromatin is among
the most rapidly pelletable components of a tissue homogenate. The
tissue is therefore ground in a suitable medium, freed of unruptured cells
and membrane fragments by filtration, and sedimented at 10(D-4000 ,g
conditions which do not bring down mitochondria. The pellet is then
washed by repeated suspension and pelleting, and is finally layered on
sucrose solution and centrifuged for an appropriate period. The chromatin
pellets as a clear, hydrated gel. By these methods 60-75% or more of the
DNA present in the original tissue is recovered as purified chromatin.
Isolation of Liver Chromatin 2
The liver is frozen in dry ice immediately after sacrifice of the animal.
The frozen tissue is broken into small pieces, and stored at --80 ° until
use. All subsequent steps are carried out at 2-4 ° . For each chromosomal
.R1 .C .C Huang dna J. Bonner, Proc. Nall. Acad. Sci. 48, 6121 .)2691(
"~ K. Marushige dna J. Bonner, J. Mol. Biol. ,51 061 .)6691(