Flexibility and distributive synthesis regulate RNA priming and handoff in human DNA polymerase α-primase

Cordoba J, Mullins E, Salay L, Eichman B, Chazin W
Journal of Molecular Biology :168330 (2023 Oct)

doi: 10.1016/j.jmb.2023.168330

Submitted to SASBDB: 2023 Jul 11
Published in SASBDB: 2023 Oct 26

SASDS87 – polymerase alpha-primase polΔcat-prim in the absence of substrate

DNA polymerase alpha subunit BDNA polymerase alpha catalytic subunitDNA primase large subunitDNA primase small subunit experimental SAS data

BILBOMD model

Sample:	DNA polymerase alpha subunit B monomer, 49 kDa Homo sapiens protein DNA polymerase alpha catalytic subunit monomer, 23 kDa Homo sapiens protein DNA primase large subunit monomer, 59 kDa Homo sapiens protein DNA primase small subunit monomer, 50 kDa Homo sapiens protein
Buffer:	20 mM HEPES, 150 mM NaCl, 5 mM MgCl2, 1 mM TCEP, pH: 7.5
Experiment:	SAXS data collected at 12.3.1 (SIBYLS), Advanced Light Source (ALS) on 2018 Dec 13

R_g^Guinier	4.6	nm
D_max	15.1	nm
Volume^Porod	300	nm³

SASDS97 – polymerase alpha-primase polΔcat-prim bound to DNA in the presence of RNA initiation factors

DNA polymerase alpha subunit BDNA polymerase alpha catalytic subunitDNA primase large subunitDNA primase small subunit22mer DNA template experimental SAS data

DNA polymerase alpha subunit B DNA polymerase alpha catalytic subunit DNA primase large subunit DNA primase small subunit 22mer DNA template Kratky plot

Sample:	DNA polymerase alpha subunit B monomer, 49 kDa Homo sapiens protein DNA polymerase alpha catalytic subunit monomer, 23 kDa Homo sapiens protein DNA primase large subunit monomer, 59 kDa Homo sapiens protein DNA primase small subunit monomer, 50 kDa Homo sapiens protein 22mer DNA template monomer, 7 kDa DNA
Buffer:	20 mM HEPES, 150 mM NaCl, 5 mM MnCl2, 1 mM TCEP, 1 mM 5'-guanylylmethylenediphosphonate (GMPCPP), pH: 7.5
Experiment:	SAXS data collected at 12.3.1 (SIBYLS), Advanced Light Source (ALS) on 2020 Dec 4

R_g^Guinier	4.6	nm
D_max	14.2	nm
Volume^Porod	310	nm³

SASDSA7 – polymerase alpha-primase polΔcat-prim bound to DNA in the presence of an RNA-primed template

DNA polymerase alpha subunit BDNA polymerase alpha catalytic subunitDNA primase large subunitDNA primase small subunit29 mer DNA template8mer RNA primer experimental SAS data

DNA polymerase alpha subunit B DNA polymerase alpha catalytic subunit DNA primase large subunit DNA primase small subunit 29 mer DNA template 8mer RNA primer Kratky plot

Sample:	DNA polymerase alpha subunit B monomer, 49 kDa Homo sapiens protein DNA polymerase alpha catalytic subunit monomer, 23 kDa Homo sapiens protein DNA primase large subunit monomer, 59 kDa Homo sapiens protein DNA primase small subunit monomer, 50 kDa Homo sapiens protein 29 mer DNA template monomer, 9 kDa DNA 8mer RNA primer monomer, 3 kDa RNA
Buffer:	20 mM HEPES, 150 mM NaCl, 5 mM MgCl2, 1 mM TCEP, pH: 7.5
Experiment:	SAXS data collected at 12.3.1 (SIBYLS), Advanced Light Source (ALS) on 2023 Mar 28

R_g^Guinier	4.9	nm
D_max	15.3	nm
Volume^Porod	344	nm³

SASDSB7 – polymerase alpha-primase polΔcat-primΔ2C in the absence of substrate

DNA polymerase alpha subunit BDNA polymerase alpha catalytic subunitDNA primase small subunitDNA primase large subunit experimental SAS data

DNA polymerase alpha subunit B DNA polymerase alpha catalytic subunit DNA primase small subunit DNA primase large subunit Kratky plot

Sample:	DNA polymerase alpha subunit B monomer, 49 kDa Homo sapiens protein DNA polymerase alpha catalytic subunit monomer, 23 kDa Homo sapiens protein DNA primase small subunit monomer, 50 kDa Homo sapiens protein DNA primase large subunit monomer, 31 kDa Homo sapiens protein
Buffer:	20 mM HEPES, 150 mM NaCl, 5 mM MgCl2, 1 mM TCEP, pH: 7.5
Experiment:	SAXS data collected at 12.3.1 (SIBYLS), Advanced Light Source (ALS) on 2019 Apr 15

R_g^Guinier	4.5	nm
D_max	14.5	nm
Volume^Porod	221	nm³