Functional tests for the characterization of surfactant protein B (SP-B) and a fluorescent SP-B analog.
Surfactant protein B (SP-B) enhances lipid insertion into the alveolar air/liquid interface upon inhalation. The aim of this study was (i) to apply a palette of tests for a detailed biochemical and biophysical characterization of SP-B and (ii) to use these tests to compare native SP-B with a fluorescent (Bodipy) SP-B analog. The method of labeling was fast and resulted in a covalent fluorophore-protein bond. The ability of both proteins to spread a surfactant film on top of a buffer surface was determined in a spreading tray using the Wilhelmy plate technique to allow detection of alterations in surface tension and calculation of spreading velocities. In a captive bubble surfactometer surface tensions of spread films were measured. Similar biophysical properties were found for both native and Bodipy-labeled SP-B. It is concluded that the combination of tests used allows detection of small differences in structure and activity between the two proteins.
Diemel
R V
RV
Department of Anaesthesiology and Critical Care Medicine, The Leopold-Franzens-University of Innsbruck, Austria. r.diemel@vet.uu.nl
Bader
D
D
Walch
M
M
Hotter
B
B
van Golde
L M
LM
Amann
A
A
Haagsman
H P
HP
Putz
G
G
D015060:1,2-Dipalmitoylphosphatidylcholine / Q000378:metabolism; D000818:Animals; D001896:Boron Compounds / Q000378:metabolism*; D018893:Bronchoalveolar Lavage; D002021:Buffers; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D005456:Fluorescent Dyes / Q000378:metabolism*; D005898:Glass; D008823:Micelles; D010713:Phosphatidylcholines / Q000378:metabolism; D010715:Phosphatidylglycerols / Q000378:metabolism; D011510:Proteolipids / Q000737:chemistry* / Q000378:metabolism*; D011663:Pulmonary Surfactants / Q000737:chemistry* / Q000378:metabolism*; D011721:Pyrenes / Q000378:metabolism; D020539:Sequence Analysis, Protein; D021241:Spectrometry, Mass, Electrospray Ionization; D013329:Structure-Activity Relationship; D013500:Surface Tension
C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D001896:Boron Compounds; D002021:Buffers; D005456:Fluorescent Dyes; D008823:Micelles; D010713:Phosphatidylcholines; D010715:Phosphatidylglycerols; D011510:Proteolipids; D011663:Pulmonary Surfactants; D011721:Pyrenes; D015060:1,2-Dipalmitoylphosphatidylcholine; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol; C030984:pyrene