Increased Electrophoretic Mobility of Long-Type GATA-6 Transcription Factor upon Substitution of Its PEST Sequence
Author(s)
Kanako Obayashi1,
Kayoko Takada1,
Kazuaki Ohashi2,
Ayako Ohashi-Kobayashi3,
Mayumi Nakanishi-Matsui4,
Makoto Araki5,
Masatomo Maeda5*
Affiliation(s)
1 Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan. 2 Department of Medical Biochemistry, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan. 3 Department of Immunobiology, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan. 4 Department of Biochemistry, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan. 5 Department of Molecular Biology, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan.
1 Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan. 2 Department of Medical Biochemistry, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan. 3 Department of Immunobiology, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan. 4 Department of Biochemistry, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan. 5 Department of Molecular Biology, School of Pharmacy, Iwate Medical University, Shiwa-Gun, Japan.
ABSTRACT
The transcriptional factor GATA-6 gene produces two translational isoforms from a single mRNA through ribosomal leaky scanning. L-type GATA-6 has an extension of 146 amino acid residues at its amino terminus. In the extension, there is a unique PEST sequence (Glu31-Cys46), which is composed of an amino terminal Pro-rich segment and a carboxyl terminal Ser-cluster. Substitution of either half of the PEST sequence with Ala residues by cassette mutagenesis reduced the apparent molecular size of L-type GATA-6 on SDS-polyacrylamide gel-electrophoresis. However, the effect of substitution of the Pro-rich segment was much more significant; the mobility increase of the Pro-rich segment on the gel was 13% while that of the Ser-cluster was 8%. Substitution of each amino acid residue demonstrated that the effect of Pro substitution is greater than that of the Ser and Thr residues. Such increased mobility of L-type GATA-6 in the presence of a detergent may apparently correlate with the decrease in transcription activity in vivo as determined by means of luciferase reporter gene assay. The activity of ΔAla (with Ala residues instead of the PEST sequence) was reduced to one fifth of that of ΔA (with the PEST sequence). These results suggest that the PEST sequence of L-type GATA-6 does not function as a constitutive protein degradation signal, but rather plays structural and functional roles in the activation of gene expression on the GATA responsive promoter.
The transcriptional factor GATA-6 gene produces two translational isoforms from a single mRNA through ribosomal leaky scanning. L-type GATA-6 has an extension of 146 amino acid residues at its amino terminus. In the extension, there is a unique PEST sequence (Glu31-Cys46), which is composed of an amino terminal Pro-rich segment and a carboxyl terminal Ser-cluster. Substitution of either half of the PEST sequence with Ala residues by cassette mutagenesis reduced the apparent molecular size of L-type GATA-6 on SDS-polyacrylamide gel-electrophoresis. However, the effect of substitution of the Pro-rich segment was much more significant; the mobility increase of the Pro-rich segment on the gel was 13% while that of the Ser-cluster was 8%. Substitution of each amino acid residue demonstrated that the effect of Pro substitution is greater than that of the Ser and Thr residues. Such increased mobility of L-type GATA-6 in the presence of a detergent may apparently correlate with the decrease in transcription activity in vivo as determined by means of luciferase reporter gene assay. The activity of ΔAla (with Ala residues instead of the PEST sequence) was reduced to one fifth of that of ΔA (with the PEST sequence). These results suggest that the PEST sequence of L-type GATA-6 does not function as a constitutive protein degradation signal, but rather plays structural and functional roles in the activation of gene expression on the GATA responsive promoter.
KEYWORDS
Cassette Mutagenesis, Long-Type GATA-6, Mobility on Gel-Electrophoresis, PEST Sequence, Proline-Rich Segment, Transcription Factor
Cassette Mutagenesis, Long-Type GATA-6, Mobility on Gel-Electrophoresis, PEST Sequence, Proline-Rich Segment, Transcription Factor
Cite this paper
Obayashi, K. , Takada, K. , Ohashi, K. , Ohashi-Kobayashi, A. , Nakanishi-Matsui, M. , Araki, M. and Maeda, M. (2014) Increased Electrophoretic Mobility of Long-Type GATA-6 Transcription Factor upon Substitution of Its PEST Sequence. Advances in Bioscience and Biotechnology, 5, 1032-1042. doi: 10.4236/abb.2014.513118.
Obayashi, K. , Takada, K. , Ohashi, K. , Ohashi-Kobayashi, A. , Nakanishi-Matsui, M. , Araki, M. and Maeda, M. (2014) Increased Electrophoretic Mobility of Long-Type GATA-6 Transcription Factor upon Substitution of Its PEST Sequence. Advances in Bioscience and Biotechnology, 5, 1032-1042. doi: 10.4236/abb.2014.513118.
References
[1] Maeda, M., Kubo, K., Nishi, T. and Futai, M. (1996) Roles of Gastric GATA DNA-Binding Proteins. The Journal of Experimental Biology, 199, 513-520. [2] Molkentin, J.D. (2000) The Zinc Finger-Containing Transcription Factors GATA-4, -5, and -6; Ubiquitously Expressed Regulators of Tissue-Specific Gene Expression. The Journal of Biological Chemistry, 275, 38949-38952.
http://dx.doi.org/10.1074/jbc.R000029200 [3] Maeda, M., Ohashi, K. and Ohashi-Kobayashi, A. (2005) Further Extension of Mammalian GATA-6. Development Growth and Differentiation, 47, 591-600.
http://dx.doi.org/10.1111/j.1440-169X.2005.00837.x [4] Ohara, Y., Atarashi, T., Ishibashi, T., Ohashi-Kobayashi, A. and Maeda, M. (2006) GATA-4 Gene Organization and Analysis of Its Promoter. Biological Pharmaceutical Bulletin, 29, 410-419.
http://dx.doi.org/10.1248/bpb.29.410 [5] Crossley, M., Merika, M. and Orkin, S.H. (1995) Self-Association of the Erythroid Transcription Factor GATA-1 Mediated by Its Zinc Finger Domain. Molecular and Cellular Biology, 15, 2448-2456. [6] Morrisey, E.E., Ip, H.S., Tang, Z. and Parmacek, M.S. (1997) GATA-4 Activates Transcription via Two Novel Domains That Are Conserved within the GATA-4/5/6 Subfamily. The Journal of Biological Chemistry, 272, 8515-8524.
http://dx.doi.org/10.1074/jbc.272.13.8515 [7] Takeda, M., Obayashi, K., Kobayashi, A. and Maeda, M. (2004) A Unique Role of an Amino Terminal 16-Residue Region of Long-Type GATA-6. Journal of Biochemstry, 135, 639-650.
http://dx.doi.org/10.1093/jb/mvh077 [8] Brewer, A., Gove, C., Davies, A., McNulty, C., Barrow, D., Koutsourakisi, M., Farzaneh, F., Pizzey, J., Bomford, A. and Patient, R. (1999) The Human and Mouse GATA-6 Genes Utilize Two Promoters and Two Initiation Codons. The Journal of Biological Chemistry, 274, 38004-38016.
http://dx.doi.org/10.1074/jbc.274.53.38004 [9] Obayashi, K., Takada, K., Ohashi, K., Kobayashi-Ohashi, A. and Maeda, M. (2012) Role of the PEST Sequence in the Long-Type GATA-6 DNA-Binding Protein Expressed in Human Cancer Cells. Advances in Bioscience and Biotechnology, 3, 314-320.
http://dx.doi.org/10.4236/abb.2012.34045 [10] Rogers, S., Wells, R. and Rechsteiner, M. (1986) Amino Acid Sequence Common to Rapidly Degraded Proteins: The PEST Hypothesis. Science, 234, 364-368.
http://dx.doi.org/10.1126/science.2876518 [11] Sanger, F., Coulson, A.R., Barrell, B.G., Smith, A.J.H. and Roe, B.A. (1980) Cloning in Single-Stranded Bacteriophage as an Aid to Rapid DNA Sequencing. Journal of Molecular Biology, 143, 161-178.
http://dx.doi.org/10.1016/0022-2836(80)90196-5 [12] Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual. 2nd Edition, Cold Spring Harbor Laboratory, Cold Spring Harbor. [13] Laemmli, U.K. (1970) Cleavage of Structural Proteins during the Assembly of the Bacteriophage T4. Nature, 227, 680-685.
http://dx.doi.org/10.1038/227680a0 [14] Tsuge, T., Uetani, K., Sato, R., Ohashi-Kobayashi, A. and Maeda, M. (2008) Cyclic AMP-Dependent Proteolysis of GATA-6 Expressed on the Intracellular Membrane. Cell Biology International, 32, 298-303.
http://dx.doi.org/10.1016/j.cellbi.2007.10.005 [15] Smith, P.K., Krohn, R.I., Hermanson, G.T., Mallia, A.K., Gartner, F.H., Provenzano, M.D., Fujimoto, E.K., Goeke, N.M., Olson, B.J. and Klenk, D.C. (1985) Measurement of Proteins Using Bicinchroninic Acid. Analytical Biochemistry, 150, 76-85.
http://dx.doi.org/10.1016/0003-2697(85)90442-7 [16] Takada, K., Obayashi, K., Ohashi, K., Kobayashi-Ohashi, A., Nakanishi-Matsui, M. and Maeda, M. (2014) Amino-Terminal Extension of 146 Residues of L-Type GATA-6 Is Required for Transcriptional Activation but Not for Self-Association. Biochemical and Biophysical Research Communications, 452, 962-966.
http://dx.doi.org/10.1016/j.bbrc.2014.09.019 [17] Kozak, M. (1991) Structural Features in Eukaryotic mRMAs That Modulate the Initiation of Translation. The Journal of Biological Chemistry, 266, 19867-19870. [18] Singh, G.P., Ganapathi, M., Sandhu, K.S. and Dash, D. (2006) Intrinsic Unstructuredness and Abundance of PEST Motifs in Eukaryotic Proteomes. PROTEINS: Structure, Function, and Bioinformatics, 62, 309-315.
http://dx.doi.org/10.1002/prot.20746 [19] Sue, S.-C. and Dyson, H.J. (2009) Interaction of the IκBα C-Terminal PEST Sequence with NF-κB: Insights into the Inhibition of NF-κB DNA Binding by IκBα. Journal of Molecular Biology, 388, 824-838.
http://dx.doi.org/10.1016/j.jmb.2009.03.048 [20] Davidson, D., Cloutier, J.-F., Gregorieff, A. and Veillette, A. (1997) Inhibitory Tyrosine Protein Kinase p50csk Is Associated with Protein-Tyrosine Phosphatase PTP-PEST in Hemopoietic and Non-Hemopoietic Cells. The Journal of Biological Chemistry, 272, 23455-23462.
http://dx.doi.org/10.1074/jbc.272.37.23455 [21] Charest, A., Wagner, J., Kwan, M. and Tremblay, M.L. (1997) Coupling of the Murine Protein Tyrosine Phosphatase PEST to the Epidermal Growth Factor (EGF) Receptor through a Src Homology 3 (SH3) Domain-Mediated Association with Grb2. Oncogene, 14, 1643-1651.
http://dx.doi.org/10.1038/sj.onc.1201008 [22] Playford, M.P., Lyons, P.D., Sastry, S.K. and Schaller, M.D. (2006) Identification of a Filamin Docking Site on PTP-PEST. The Journal of Biological Chemistry, 281, 34104-34112.
http://dx.doi.org/10.1074/jbc.M606277200 [23] Kiran, M. and Nagarajaram, H.A. (2013) Global versus Local Hubs in Human Protein-Protein Interaction Network. Journal of Proteome Research, 12, 5436-5446.
http://dx.doi.org/10.1021/pr4002788 [24] Adzhubei, A.A., Sternberg, M.J.E. and Makarov, A.A. (2013) Polyproline-II Helix in Proteins: Structure and Function. Journal of Molecular Biology, 425, 2100-2132.
http://dx.doi.org/10.1016/j.jmb.2013.03.018 [25] Dornan, D., Shimizu, H., Burch, L., Smith, A.J. and Hupp, T.R. (2003) The Proline Repeat Domain of p53 Binds Directly to the Transcriptional Coactivator p300 and Allosterically Controls DNA-Dependent Acetylation of p53. Molecular and Cellular Biology, 23, 8846-8861.
http://dx.doi.org/10.1128/MCB.23.23.8846-8861.2003 [26] Singh, V., Lin, R., Yang, J., Cha, B., Sarker, R., Tse, C.M. and Donowitz, M. (2014) AKT and GSK-3 Are Necessary for Direct Ezrin Binding to NHE3 as Part of a C-Terminal Stimulatory Complex: Role of a Novel Ser-Rich NHE3 C-Terminal Motif in NHE3 Activity and Trafficking. The Journal of Biological Chemistry, 289, 5449-5461.
http://dx.doi.org/10.1074/jbc.M113.521336 [27] Zheng, H., You, H., Zhou, X.Z., Murray, S.A., Uchida, T., Wulf, G., Gu, L., Tang, X., Lu, K.P. and Xiao, Z.-X.J. (2002) The Prolylisomerase Pin1 Is a Regulator of p53 in Genotoxic Response. Nature, 419, 849-853.
http://dx.doi.org/10.1038/nature01116 [28] Pham, D.Q.-D. and Sivasubramanian, N. (1992) Sequence and in Vitro Translational Analysis of a 1629-Nucleotide ORF in Autographa californica Nuclear Polyhedrosis Virus Strain E2. Gene, 122, 345-348.
http://dx.doi.org/10.1016/0378-1119(92)90224-D [29] John, M.E. and Keller, G. (1995) Characterization of mRNA for a Proline-Rich Protein of Cotton Fiber. Plant Physiology, 108, 669-676.
http://dx.doi.org/10.1104/pp.108.2.669 [30] Rath, A., Glibowicka, M., Nadeau, V.G., Chen, G. and Deber, C.M. (2009) Detergent Binding Explains Anomalous SDS-PAGE Migration of Membrane Proteins. Proceedings of the National Academy of Sciences of the United States of America, 106, 1760-1765.
http://dx.doi.org/10.1073/pnas.0813167106 [31] Shi, Y., Mowery, R.A., Ashley, J., Hentz, M., Ramirez, A.J., Bilgicer, B., Slunt-Brown, H., Borchelt, D.R. and Shaw, B.F. (2012) Abnormal SDS-PAGE Migration of Cytosolic Proteins Can Identify Domains and Mechanisms That Control Surfactant Binding. The Protein Society, 21, 1197-1209.
http://dx.doi.org/10.1002/pro.2107
[1] Maeda, M., Kubo, K., Nishi, T. and Futai, M. (1996) Roles of Gastric GATA DNA-Binding Proteins. The Journal of Experimental Biology, 199, 513-520. [2] Molkentin, J.D. (2000) The Zinc Finger-Containing Transcription Factors GATA-4, -5, and -6; Ubiquitously Expressed Regulators of Tissue-Specific Gene Expression. The Journal of Biological Chemistry, 275, 38949-38952.
http://dx.doi.org/10.1074/jbc.R000029200 [3] Maeda, M., Ohashi, K. and Ohashi-Kobayashi, A. (2005) Further Extension of Mammalian GATA-6. Development Growth and Differentiation, 47, 591-600.
http://dx.doi.org/10.1111/j.1440-169X.2005.00837.x [4] Ohara, Y., Atarashi, T., Ishibashi, T., Ohashi-Kobayashi, A. and Maeda, M. (2006) GATA-4 Gene Organization and Analysis of Its Promoter. Biological Pharmaceutical Bulletin, 29, 410-419.
http://dx.doi.org/10.1248/bpb.29.410 [5] Crossley, M., Merika, M. and Orkin, S.H. (1995) Self-Association of the Erythroid Transcription Factor GATA-1 Mediated by Its Zinc Finger Domain. Molecular and Cellular Biology, 15, 2448-2456. [6] Morrisey, E.E., Ip, H.S., Tang, Z. and Parmacek, M.S. (1997) GATA-4 Activates Transcription via Two Novel Domains That Are Conserved within the GATA-4/5/6 Subfamily. The Journal of Biological Chemistry, 272, 8515-8524.
http://dx.doi.org/10.1074/jbc.272.13.8515 [7] Takeda, M., Obayashi, K., Kobayashi, A. and Maeda, M. (2004) A Unique Role of an Amino Terminal 16-Residue Region of Long-Type GATA-6. Journal of Biochemstry, 135, 639-650.
http://dx.doi.org/10.1093/jb/mvh077 [8] Brewer, A., Gove, C., Davies, A., McNulty, C., Barrow, D., Koutsourakisi, M., Farzaneh, F., Pizzey, J., Bomford, A. and Patient, R. (1999) The Human and Mouse GATA-6 Genes Utilize Two Promoters and Two Initiation Codons. The Journal of Biological Chemistry, 274, 38004-38016.
http://dx.doi.org/10.1074/jbc.274.53.38004 [9] Obayashi, K., Takada, K., Ohashi, K., Kobayashi-Ohashi, A. and Maeda, M. (2012) Role of the PEST Sequence in the Long-Type GATA-6 DNA-Binding Protein Expressed in Human Cancer Cells. Advances in Bioscience and Biotechnology, 3, 314-320.
http://dx.doi.org/10.4236/abb.2012.34045 [10] Rogers, S., Wells, R. and Rechsteiner, M. (1986) Amino Acid Sequence Common to Rapidly Degraded Proteins: The PEST Hypothesis. Science, 234, 364-368.
http://dx.doi.org/10.1126/science.2876518 [11] Sanger, F., Coulson, A.R., Barrell, B.G., Smith, A.J.H. and Roe, B.A. (1980) Cloning in Single-Stranded Bacteriophage as an Aid to Rapid DNA Sequencing. Journal of Molecular Biology, 143, 161-178.
http://dx.doi.org/10.1016/0022-2836(80)90196-5 [12] Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual. 2nd Edition, Cold Spring Harbor Laboratory, Cold Spring Harbor. [13] Laemmli, U.K. (1970) Cleavage of Structural Proteins during the Assembly of the Bacteriophage T4. Nature, 227, 680-685.
http://dx.doi.org/10.1038/227680a0 [14] Tsuge, T., Uetani, K., Sato, R., Ohashi-Kobayashi, A. and Maeda, M. (2008) Cyclic AMP-Dependent Proteolysis of GATA-6 Expressed on the Intracellular Membrane. Cell Biology International, 32, 298-303.
http://dx.doi.org/10.1016/j.cellbi.2007.10.005 [15] Smith, P.K., Krohn, R.I., Hermanson, G.T., Mallia, A.K., Gartner, F.H., Provenzano, M.D., Fujimoto, E.K., Goeke, N.M., Olson, B.J. and Klenk, D.C. (1985) Measurement of Proteins Using Bicinchroninic Acid. Analytical Biochemistry, 150, 76-85.
http://dx.doi.org/10.1016/0003-2697(85)90442-7 [16] Takada, K., Obayashi, K., Ohashi, K., Kobayashi-Ohashi, A., Nakanishi-Matsui, M. and Maeda, M. (2014) Amino-Terminal Extension of 146 Residues of L-Type GATA-6 Is Required for Transcriptional Activation but Not for Self-Association. Biochemical and Biophysical Research Communications, 452, 962-966.
http://dx.doi.org/10.1016/j.bbrc.2014.09.019 [17] Kozak, M. (1991) Structural Features in Eukaryotic mRMAs That Modulate the Initiation of Translation. The Journal of Biological Chemistry, 266, 19867-19870. [18] Singh, G.P., Ganapathi, M., Sandhu, K.S. and Dash, D. (2006) Intrinsic Unstructuredness and Abundance of PEST Motifs in Eukaryotic Proteomes. PROTEINS: Structure, Function, and Bioinformatics, 62, 309-315.
http://dx.doi.org/10.1002/prot.20746 [19] Sue, S.-C. and Dyson, H.J. (2009) Interaction of the IκBα C-Terminal PEST Sequence with NF-κB: Insights into the Inhibition of NF-κB DNA Binding by IκBα. Journal of Molecular Biology, 388, 824-838.
http://dx.doi.org/10.1016/j.jmb.2009.03.048 [20] Davidson, D., Cloutier, J.-F., Gregorieff, A. and Veillette, A. (1997) Inhibitory Tyrosine Protein Kinase p50csk Is Associated with Protein-Tyrosine Phosphatase PTP-PEST in Hemopoietic and Non-Hemopoietic Cells. The Journal of Biological Chemistry, 272, 23455-23462.
http://dx.doi.org/10.1074/jbc.272.37.23455 [21] Charest, A., Wagner, J., Kwan, M. and Tremblay, M.L. (1997) Coupling of the Murine Protein Tyrosine Phosphatase PEST to the Epidermal Growth Factor (EGF) Receptor through a Src Homology 3 (SH3) Domain-Mediated Association with Grb2. Oncogene, 14, 1643-1651.
http://dx.doi.org/10.1038/sj.onc.1201008 [22] Playford, M.P., Lyons, P.D., Sastry, S.K. and Schaller, M.D. (2006) Identification of a Filamin Docking Site on PTP-PEST. The Journal of Biological Chemistry, 281, 34104-34112.
http://dx.doi.org/10.1074/jbc.M606277200 [23] Kiran, M. and Nagarajaram, H.A. (2013) Global versus Local Hubs in Human Protein-Protein Interaction Network. Journal of Proteome Research, 12, 5436-5446.
http://dx.doi.org/10.1021/pr4002788 [24] Adzhubei, A.A., Sternberg, M.J.E. and Makarov, A.A. (2013) Polyproline-II Helix in Proteins: Structure and Function. Journal of Molecular Biology, 425, 2100-2132.
http://dx.doi.org/10.1016/j.jmb.2013.03.018 [25] Dornan, D., Shimizu, H., Burch, L., Smith, A.J. and Hupp, T.R. (2003) The Proline Repeat Domain of p53 Binds Directly to the Transcriptional Coactivator p300 and Allosterically Controls DNA-Dependent Acetylation of p53. Molecular and Cellular Biology, 23, 8846-8861.
http://dx.doi.org/10.1128/MCB.23.23.8846-8861.2003 [26] Singh, V., Lin, R., Yang, J., Cha, B., Sarker, R., Tse, C.M. and Donowitz, M. (2014) AKT and GSK-3 Are Necessary for Direct Ezrin Binding to NHE3 as Part of a C-Terminal Stimulatory Complex: Role of a Novel Ser-Rich NHE3 C-Terminal Motif in NHE3 Activity and Trafficking. The Journal of Biological Chemistry, 289, 5449-5461.
http://dx.doi.org/10.1074/jbc.M113.521336 [27] Zheng, H., You, H., Zhou, X.Z., Murray, S.A., Uchida, T., Wulf, G., Gu, L., Tang, X., Lu, K.P. and Xiao, Z.-X.J. (2002) The Prolylisomerase Pin1 Is a Regulator of p53 in Genotoxic Response. Nature, 419, 849-853.
http://dx.doi.org/10.1038/nature01116 [28] Pham, D.Q.-D. and Sivasubramanian, N. (1992) Sequence and in Vitro Translational Analysis of a 1629-Nucleotide ORF in Autographa californica Nuclear Polyhedrosis Virus Strain E2. Gene, 122, 345-348.
http://dx.doi.org/10.1016/0378-1119(92)90224-D [29] John, M.E. and Keller, G. (1995) Characterization of mRNA for a Proline-Rich Protein of Cotton Fiber. Plant Physiology, 108, 669-676.
http://dx.doi.org/10.1104/pp.108.2.669 [30] Rath, A., Glibowicka, M., Nadeau, V.G., Chen, G. and Deber, C.M. (2009) Detergent Binding Explains Anomalous SDS-PAGE Migration of Membrane Proteins. Proceedings of the National Academy of Sciences of the United States of America, 106, 1760-1765.
http://dx.doi.org/10.1073/pnas.0813167106 [31] Shi, Y., Mowery, R.A., Ashley, J., Hentz, M., Ramirez, A.J., Bilgicer, B., Slunt-Brown, H., Borchelt, D.R. and Shaw, B.F. (2012) Abnormal SDS-PAGE Migration of Cytosolic Proteins Can Identify Domains and Mechanisms That Control Surfactant Binding. The Protein Society, 21, 1197-1209.
http://dx.doi.org/10.1002/pro.2107